neuropeptide-y and Hypoglycemia

Topics: Adrenocorticotropic Hormone; Animals; Cells, Cultured; Hypoglycemia; Hypothalamo-Hypophyseal System; Insulin; Male; Neuropeptide Y; Rats; Rats, Inbred Strains; Shock, Septic; Stress, Physiological

The NPY secretory pattern after an insulin tolerance test (ITT) (0.15 IU/kg body weight) was evaluated in 8 normal men. They were infused with normal saline (control test), glucose or fructose. Insulin-induced hypoglycemia produced a significant increment in serum NPY in the control test. The infusion of fructose was unable to change the NPY secretory pattern during insulin-induced hypoglycemia. In contrast, the NPY increase during ITT was completely abolished when the concomitant infusion of glucose prevented insulin-induced hypoglycemia. These results exclude a direct role of hyperinsulinemia in the mechanism underlying the stimulation of NPY secretion during ITT. Furthermore, since glucose but not fructose crosses the blood-brain-barrier (BBB), the NPY increase during ITT appears to be generated by low glucose concentrations at the level of glucosensitive areas located inside the brain.

Topics: Adult; Blood-Brain Barrier; Brain; Fructose; Glucose; Humans; Hypoglycemia; Insulin; Male; Neuropeptide Y; Receptors, Cell Surface

1. In postganglionic sympathetic neurones and adrenal chromaffin cells, catecholamines are co-stored in vesicles with soluble peptides, including chromogranin A (CgA) and neuropeptide Y (NPY), which are subject to exocytotic co-release with catecholamines. 2. Plasma catecholamine, CgA and NPY responses to stimulators and inhibitors of sympatho-adrenal catecholamine storage and release were measured in humans. Short-term, high-intensity dynamic exercise, prolonged low-intensity dynamic exercise, and assumption of the upright posture, in decreasing order of potency, predominantly stimulated noradrenaline (NA) release from sympathetic nerve endings. Only high-intensity exercise elevated CgA and NPY, which did not peak until 2 min after exercise cessation. Stimulated NA correlated with plasma CgA 2 min after exercise, and with NPY 5 min after exercise. 3. Insulin-evoked hypoglycaemia and caffeine ingestion, in decreasing order of potency, predominantly stimulated adrenaline (AD) release from the adrenal medulla. During insulin hypoglycaemia AD and CgA rose, but NPY was unchanged. Neither NPY nor CgA were altered by caffeine. The rise in CgA after intense adrenal medullary stimulation was greater than its rise after intense sympathetic neuronal stimulation (1.4-versus 1.2-fold, respectively). 4. Infusion of tyramine, which disrupts sympathetic neuronal vesicular NA storage, elevated systolic blood pressure and NA, while NPY and CgA were unchanged. After reserpine, another disruptor of neuronal NA storage, NA transiently rose and then fell; NPY and CgA were unaltered. After the non-exocytotic adrenal medullary secretory stimulus glucagon. AD rose while NA, CgA and NPY did not change. After amantadine, an inhibitor of protein endocytosis, both CgA and fibrinogen rose, while NA and NPY remained unaltered. Neither CgA, NPY, nor catecholamines were altered by the catecholamine uptake and catabolism inhibitors desipramine, cortisol, and pargyline. 5. Human sympathetic nerve contained a far higher ratio of NPY to catecholamines than human adrenal medulla, while adrenal medulla contained far more CgA than sympathetic nerve. 6. It is concluded that peptides are differentially co-stored with catecholamines, with greater abundance of CgA in the adrenal medulla and NPY in sympathetic nerve. Activation of catecholamine release from either the adrenal medulla or sympathetic nerves, therefore, results in quite different changes in plasma concentrations of the catecholamine s

Topics: Adrenal Medulla; Adult; Aged; Amantadine; Analysis of Variance; Caffeine; Chromogranin A; Chromogranins; Endocytosis; Exercise; Female; Humans; Hypoglycemia; Male; Middle Aged; Neuropeptide Y; Norepinephrine; Posture; Tyramine; Vascular Resistance

Hindbrain energy state shapes hypothalamic control of glucostasis. Dorsal vagal complex (DVC) L-lactate deficiency is a potent glucose-stimulatory signal that triggers neuronal transcriptional activation in key hypothalamic metabolic loci. The energy gauge AMPK is activated in DVC metabolic-sensory A2 noradrenergic neurons by hypoglycemia-associated lactoprivation, but sensor reactivity is diminished by antecedent hypoglycemia (AH). Current research addressed the premise that AH alters hindbrain lactoprivic regulation of hypothalamic metabolic transmitter function. AH did not modify reductions in A2 dopamine-beta-hydroxylase and monocarboxylate-2 (MCT2) protein expression elicited by caudal fourth ventricular delivery of the MCT inhibitor alpha-cyano-4-hydroxycinnamic acid (4CIN), but attenuated 4CIN activation of A2 AMPK. 4CIN constraint of hypothalamic norepinephrine (NE) activity was averted by AH in a site-specific manner. 4CIN induction of Fos immunolabeling in hypothalamic arcuate (ARH), ventromedial (VMN), dorsomedial (DMN) and paraventricular (PVN) nuclei and lateral hypothalamic area (LHA) was avoided by AH. AH affected reactivity of select hypothalamic metabolic neurotransmitter/enzyme marker proteins, e.g. ARH neuropeptide Y, VMN glutamate decarboxylase, DMN RFamide-related peptide-1 and -3, and LHA orexin-A profiles to 4CIN, but did not alleviate drug inhibition of ARH proopiomelanocortin. AH prevented 4CIN augmentation of circulating glucagon, but did not alter hyperglycemic or hypocorticosteronemic responses to that treatment. Results identify hindbrain lactate deficiency as a stimulus for glucagon secretion, and imply that habituation of this critical counter-regulatory hormone to recurring hypoglycemia may involve one or more hypothalamic neurotransmitters characterized here by acclimation to this critical sensory stimulus.

Topics: Animals; Blood Glucose; Hypoglycemia; Hypothalamus; Insulin; Male; Neurons; Neuropeptide Y; Norepinephrine; Pro-Opiomelanocortin; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Rhombencephalon; Transcriptional Activation

Pharmacologic activation of the hindbrain dorsal vagal complex energy sensor 5'-adenosine monophosphate-activated protein kinase (AMPK) causes site-specific adjustments in hypothalamic AMPK activity. DVC A2 noradrenergic neurons are a likely source of metabolo-sensory cues to downstream network components as they express substrate fuel-sensitive AMPK. This study investigated the hypothesis that DVC AMPK controls hypothalamic sensor, metabolic effector transmitter, and counter-regulatory hormone responses to insulin-induced hypoglycemia. Male rats were injected into the caudal fourth ventricle with the AMPK inhibitor compound C (Ccor vehicle before hypoglycemia. Arcuate (ARH), ventromedial (VMN), and dorsomedial (DMN) nuclei and lateral hypothalamic area (LHA) were micropunch-dissected for norepinephrine ELISA and Western blot analyses. Hypoglycemic stimulation of norepinephrine activity in each site was impeded by compound C. Hypoglycemia caused drug-revocable (ARH) or -refractory (VMN, DMN) reductions in AMPK, alongside hindbrain AMPK-dependent augmentation of phospho-AMPK expression in each location. Compound C prevented hypoglycemic augmentation of gluco-stimulatory ARH neuropeptide Y, VMN neuronal nitric oxide synthase, and LHA orexin-A expression, while hypoglycemic suppression of the catabolic neuron protein markers ARH pro-opiomelanocortin and VMN glutamate decarboxylase

Topics: Adrenergic Neurons; AMP-Activated Protein Kinases; Animals; Hypoglycemia; Hypothalamic Area, Lateral; Hypothalamus; Male; Neuropeptide Y; Neuropeptides; Nitric Oxide Synthase Type I; Norepinephrine; Orexins; Rats; Rats, Sprague-Dawley; Rhombencephalon; Solitary Nucleus; Vagus Nerve

Glucose is a major fuel for the central nervous system and hypoglycemia is a significant homeostatic stressor, which elicits counterregulatory reactions. Hypothalamic metabolic- and stress-related neurons initiate these actions, however recruitment of glia in control such adaptive circuit remain unknown. Groups of fed- and fasted-, vehicle-injected, and fasted + insulin-injected male mice were compared in this study. Bolus insulin administration to fasted mice resulted in hypoglycemia, which increased hypothalamo-pituitary-adrenal (HPA) axis- and sympathetic activity, increased transcription of neuropeptide Y (Npy) and agouti-related peptide (Agrp) in the hypothalamic arcuate nucleus and activated IBA1+ microglia in the hypothalamus. Activated microglia were found in close apposition to hypoglycemia-responsive NPY neurons. Inhibition of microglia by minocycline increased counterregulatory sympathetic response to hypoglycemia. Fractalkine-CX3CR1 signaling plays a role in control of microglia during hypoglycemia, because density and solidity of IBA1-ir profiles was attenuated in fasted, insulin-treated, CX3CR1 KO mice, which was parallel with exaggerated neuropeptide responses and higher blood glucose levels following insulin administration. Hypoglycemia increased Il-1b expression in the arcuate nucleus, while IL-1a/b knockout mice display improved glycemic control to insulin administration. In conclusion, activated microglia in the arcuate nucleus interferes with central counterregulatory responses to hypoglycemia. These results underscore involvement of microglia in hypothalamic regulation of glucose homeostasis.

Topics: Agouti-Related Protein; Animals; Arcuate Nucleus of Hypothalamus; Blood Glucose; Fasting; Homeostasis; Hypoglycemia; Hypothalamo-Hypophyseal System; Insulin; Interleukin-1alpha; Interleukin-1beta; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Microglia; Neurons; Neuropeptide Y; Peptide Fragments

The hypothalamic energy sensor adenosine 5'-monophosphate-activated protein kinase (AMPK), an important regulator of counter-regulatory responses to hypoglycemia, responds to pharmacological manipulation of hindbrain AMPK activity. Dorsomedial hindbrain A2 noradrenergic neurons express hypoglycemia-sensitive metabolo-sensory biomarkers, including AMPK. Here, adult male rats were pretreated by intra-caudal fourth ventricular administration of the selective neurotoxin 6-hydroxydopamine (6-OHDA) to determine if catecholamine signaling from the aforesaid site governs hypothalamic AMPK activation during insulin-induced hypoglycemia (IIH). Micropunched arcuate (ARH), ventromedial (VMH), paraventricular (PVH), dorsomedial (DMH) nuclei and lateral hypothalamic area (LHA) tissues were obtained at the neutral protamine Hagedorn insulin-induced hypoglycemic nadir, coincident with A2 AMPK activation, for Western blot analysis of AMPK, phospho-AMPK (pAMPK), and relevant metabolic neuropeptides. ARH, VMH, LHA, and DMH norepinephrine levels were altered according to insulin dose; 6-OHDA-mediated reversal of these responses was site-specific. IIH elevated LHA and reduced VMH pAMPK protein, profiles that were respectively unchanged or increased by 6-OHDA. PVH and ARH pAMPK was resistant to IIH, but augmented in ARH of neurotoxin- plus insulin-treated rats. ARH neuropeptide Y (NPY) and pro-opiomelanocortin (POMC) proteins were correspondingly increased or refractory to IIH; 6-OHDA pretreatment normalized NPY and elevated POMC expression after insulin injection. Results demonstrate site-specific bi-directional adjustments in hypothalamic AMPK reactivity to hypoglycemia. Intensification of ARH/VMH pAMPK by 6-OHDA implies dorsomedial hindbrain improvement of energy balance in those sites during IIH. Neurotoxin-mediated augmentation versus suppression of basal catabolic (ARH POMC/VMH steroidogenic factor-1) or IIH-associated anabolic (ARH NPY) neuropeptide profiles, respectively, may involve local AMPK-dependent against independent mechanisms.

Topics: Adenylate Kinase; Adrenergic Agents; Animals; Catecholamines; Hypoglycemia; Hypothalamus; Insulin; Male; Neuropeptide Y; Oxidopamine; Phosphorylation; Pro-Opiomelanocortin; Rats, Sprague-Dawley; Rhombencephalon; Signal Transduction

The mechanisms responsible for the development of the impaired awareness of hypoglycemia often seen in insulin-treated patients with diabetes remain uncertain, but cerebral adaptations to recurrent hypoglycemia are frequently hypothesized. In this issue of the JCI, Ma et al. demonstrate that neuropeptide Y (NPY) secretion from adrenal chromaffin cells persists during exposure to recurrent hypoglycemia and activation of the sympathetic nerves at the same time that epinephrine secretion is reduced. This results in the inhibition of tyrosine hydroxylase, the rate-limiting enzyme for catecholamine synthesis. These observations suggest that a peripheral mechanism downstream from the brain contributes to the development of impaired awareness of hypoglycemia.

Topics: Brain; Humans; Hypoglycemia; Insulin; Neuropeptide Y; Tyrosine 3-Monooxygenase

Hypoglycemia activates the counterregulatory response (CRR), a neural-endocrine reflex that restores euglycemia. Although effective if occasionally activated, repeated induction of the CRR leads to a decline in responsiveness and prolonged exposure to hypoglycemia. The mechanism underlying this impairment is not known. We found that the reduction in epinephrine release that characterizes a suppressed CRR involves a long-lasting form of sympatho-adrenal synaptic plasticity. Using optogenetically evoked catecholamine release, we show that recurrent hypoglycemia reduced the secretory capacity of mouse adrenal chromaffin cells. Single activation of the CRR increased the adrenal levels of tyrosine hydroxylase (TH), the rate-limiting enzyme for catecholamine synthesis, but this was prevented by repeated activation. In contrast, the level of neuropeptide Y (NPY), an adrenal cotransmitter, remained elevated after recurrent hypoglycemia. Inhibition of NPY or Y1 signaling, either transgenically or pharmacologically, prevented the attenuation of both TH expression and epinephrine release. These results indicate that impairment of the CRR involves suppressed activity at the adrenal level. Interfering with the peripheral NPY-dependent negative feedback loop may provide a way to avoid the pathophysiological consequences of recurrent hypoglycemia which are common in the diabetic state.

Topics: Adrenal Glands; Animals; Catecholamines; Chromaffin Cells; Epinephrine; Feedback, Physiological; Female; Hypoglycemia; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neuronal Plasticity; Neuropeptide Y; Recurrence; Tyrosine 3-Monooxygenase

The suprachiasmatic nucleus (SCN) and arcuate nucleus (ARC) have reciprocal connections; catabolic metabolic information activates the ARC and inhibits SCN neuronal activity. Little is known about the influence of the SCN on the ARC. Here, we investigated whether the SCN modulated the sensitivity of the ARC to catabolic metabolic conditions. ARC neuronal activity, as determined by c-Fos immunoreactivity, was increased after a hypoglycemic stimulus by 2-deoxyglucose (2DG). The highest ARC neuronal activity after 2DG was found at the end of the light period (zeitgeber 11, ZT11) with a lower activity in the beginning of the light period (zeitgeber 2, ZT2), suggesting the involvement of the SCN. The higher activation of ARC neurons after 2DG at ZT11 was associated with higher 2DG induced blood glucose levels as compared with ZT2. Unilateral SCN-lesioned animals, gave a mainly ipsilateral activation of ARC neurons at the lesioned side, suggesting an inhibitory role of the SCN on ARC neurons. The 2DG-induced counterregulatory glucose response correlated with increased ARC neuronal activity and was significantly higher in unilateral SCN-lesioned animals. Finally, the ARC as site where 2DG may, at least partly, induce a counterregulatory response was confirmed by local microdialysis of 2DG. 2DG administration in the ARC produced a higher increase in circulating glucose compared with 2DG administration in surrounding areas such as the ventromedial nucleus of the hypothalamus (VMH). We conclude that the SCN uses neuronal pathways to the ARC to gate sensory metabolic information to the brain, regulating ARC glucose sensitivity and counterregulatory responses to hypoglycemic conditions.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Fasting; Hypoglycemia; Male; Melanocyte-Stimulating Hormones; Neurons; Neuropeptide Y; Rats, Wistar; Suprachiasmatic Nucleus

Hypoglycemia elicits physiological and behavioral responses which are mediated in part by neurons within the ventrolateral medulla (VLM). The present study describes the neurochemistry of neurons activated by glucoprivation (2-deoxy-D-glucose, 2DG), specifically those within regions containing the A1, caudal C1 (cC1) and rostral C1 (rC1) cell groups. 2DG induced c-Fos immunoreactivity throughout the VLM. Activated neurons expressing prepro-cocaine and amphetamine-regulated transcript (PPCART), neuropeptide Y (NPY), glutamic acid decarboxylase (GAD67) or prepro-enkephalin (PPE) mRNA and/or immunoreactivity (-ir) for tyrosine hydroxylase (TH) were identified. TH(+) neurons were recruited in a dose-dependent manner. At high doses of 2DG [400 mg/kg, (n = 6)], 76 ± 1.2 % of activated neurons were TH(+) representing 52 ± 1.3 % of the total TH population. Virtually all activated neurons in the A1 and cC1 regions but only 60 % in the rC1 region were TH(+). Within the A1 region, TH(+), TH(+)NPY(+) and TH(+)NPY(+)PPE(+) subpopulations were activated and likely regulate vasopressin, oxytocin, and corticotrophin releasing hormone (CRH) from the hypothalamus. Within the cC1 region, non-TH neurons, TH(+)NPY(+), TH(+)NPY(+)PPCART(+), and TH(+)NPY(+)PPE(+) subpopulations were activated, likely regulating autonomic hypothalamic neurons or CRH and thyrotropin releasing hormone secretion. Within the rC1 region, non-TH neurons (40 % of those activated) were predominantly PPE(+) and were recruited by higher 2DG doses. Of the TH(+) activated neurons in the rC1 region, many expressed PPCART and half expressed NPY. The activated spinally projecting population was almost entirely TH(+)PPCART(+) and is likely to regulate adrenaline and glucagon release. These data indicate that glucoprivation activates at least nine phenotypically distinct populations of neurons in the VLM.

Topics: Animals; Blood Glucose; Cell Count; Cholera Toxin; Deoxyglucose; Disease Models, Animal; Gene Expression Regulation; Glutamate Decarboxylase; Hypoglycemia; Male; Medulla Oblongata; Nerve Tissue Proteins; Neurons; Neuropeptide Y; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; RNA, Messenger; Tyrosine 3-Monooxygenase

Nerve cell metabolic activity is monitored in multiple brain regions, including the hypothalamus and hindbrain dorsal vagal complex (DVC), but it is unclear if individual metabolosensory loci operate autonomously or interact to coordinate central nervous system (CNS) reactivity to energy imbalance. This research addressed the hypothesis that hypoglycemia-associated DVC lactoprivation stimulates hypothalamic AMPK activity and metabolic neurotransmitter expression. As DVC catecholaminergic neurons express biomarkers for metabolic monitoring, we investigated whether these cells are a source of lactate deficit signaling to the hypothalamus. Caudal fourth ventricle (CV4) infusion of the glucose metabolite l-lactate during insulin-induced hypoglycemia reversed changes in DVC A2 noradrenergic, arcuate neuropeptide Y (NPY) and pro-opiomelanocortin (POMC), and lateral hypothalamic orexin-A (ORX) neuronal AMPK activity, coincident with exacerbation of hypoglycemia. Hindbrain lactate repletion also blunted hypoglycemic upregulation of arcuate NPY mRNA and protein. This treatment did not alter hypoglycemic paraventricular oxytocin (OT) and lateral hypothalamic ORX mRNA profiles, but exacerbated or reversed adjustments in OT and ORX neuropeptide synthesis, respectively. CV4 delivery of the monocarboxylate transporter inhibitor, 4-CIN, increased A2 phosphoAMPK (pAMPK), elevated circulating glucose, and stimulated feeding, responses that were attenuated by 6-hydroxydopamine pretreatment. 4-CIN-infused rats exhibited increased (NPY, ORX neurons) or decreased (POMC neurons) pAMPK concurrent with hyperglycemia. These data show that hindbrain lactoprivic signaling regulates hypothalamic AMPK and key effector neurotransmitter responses to hypoglycemia. Evidence that A2 AMPK activity is lactate-dependent, and that DVC catecholamine cells are critical for lactoprivic control of glucose, feeding, and hypothalamic AMPK, implies A2 derivation of this metabolic regulatory stimulus.

Topics: Adrenergic Neurons; AMP-Activated Protein Kinases; Animals; Disease Models, Animal; Energy Metabolism; Enzyme Activation; Feeding Behavior; Gene Expression Regulation; Hypoglycemia; Hypothalamus; Infusions, Intraventricular; Insulin; Intracellular Signaling Peptides and Proteins; Lactic Acid; Male; Neuropeptide Y; Neuropeptides; Orexins; Pro-Opiomelanocortin; Rats; Rats, Sprague-Dawley; Rhombencephalon; RNA, Messenger; Signal Transduction

Chickens selected for low (LWS) or high (HWS) body weight for more than 56 generations now have a 10-fold difference in body weight at 56 days of age and correlated responses in appetite and glucose regulation. The LWS chickens are lean and some are anorexic, while the HWS are compulsive feeders and have a different threshold sensitivity of food intake and blood glucose to both central and peripheral insulin, respectively. We previously demonstrated that at 90-days of age, insulin-induced hypoglycemia was associated with reduced glucose transporter expression in the liver of both lines, and differences in expression of neuropeptide Y (NPY) and NPY receptor sub-type genes between LWS and HWS in the hypothalamus. The objective of this study was to determine effects of insulin-induced hypoglycemia on gene expression in the hypothalamus and liver of early post-hatch LWS and HWS chicks. On day 5 post-hatch chicks from each line were fasted for 3h and injected intraperitoneally with insulin or vehicle. At 1h post-injection, chicks were euthanized, blood glucose was measured, and hypothalamus and liver were removed. Total RNA was isolated and real time PCR performed. Insulin injection was associated with a more pronounced reduction in blood glucose in HWS compared with LWS chicks (two-way interaction; P<0.05). Aromatic L-amino acid decarboxylase, NPY, and NPY receptor sub-types 2 and 5 mRNA quantities were greater in LWS than HWS chicks in the hypothalamus (P<0.05), whereas pro-opiomelanocortin mRNA was greater in the hypothalamus of HWS than LWS (P<0.05). In the liver, glucose transporter 1, 2 and 3 (GLUT 1, 2 and 3, respectively) mRNA abundance was greater in HWS than LWS chicks (P<0.05). Compared to the vehicle, insulin treatment was associated with an increase in tryptophan hydroxylase 2 mRNA in the hypothalamus of both lines (P=0.02). In the liver of both lines, insulin treatment was associated with decreased (P=0.01) GLUT2 mRNA and increased (P=0.01) GLUT1 mRNA, compared to vehicle-treated chicks. Results suggest that NPY-associated factors and glucose transporters are differentially-expressed between LWS and HWS chickens and that HWS chicks display greater sensitivity to exogenous insulin during the early post-hatch period.

Topics: Animals; Blood Glucose; Body Weight; Chickens; Female; Gene Expression Regulation; Glucose; Glucose Transport Proteins, Facilitative; Hypoglycemia; Hypothalamus; Insulin; Liver; Neuropeptide Y; RNA, Messenger

The hypothalamic neurochemicals neuropeptide Y (NPY), orexin-A (ORX), and oxytocin (OXY) exert glucoregulatory effects upon intracerebral administration, findings that support their potential function within neural pathways that maintain glucostasis. Current understanding of how these neurotransmitter systems respond to the diabetes mellitus complication, insulin-induced hypoglycemia, is limited to knowledge of neuropeptide gene transcriptional reactivity. We investigated the hypothesis that hypoglycemia elicits hypothalamic site-specific alterations in levels of these neurochemicals, and that adjustments in local neurotransmitter availability may be regulated by catecholaminergic (CA) input from the caudal dorsomedial hindbrain. The arcuate (ARH) and paraventricular (PVH) hypothalamic nuclei and lateral hypothalamic area (LHA) were each microdissected from adult male rats pretreated by caudal fourth ventricular administration of the selective CA neurotoxin, 6-hydroxydopamine (6-OHDA), or vehicle prior to insulin (INS)-induced hypoglycemia. Hypoglycemia stimulated ARH NPY gene expression and NPY accumulation in the ARH and LHA, but not PVH. 6-OHDA pretreatment did not modify the positive NPY mRNA response to INS, but blunted hypoglycemic augmentation of ARH and LHA NPY content while increasing PVH NPY levels in response to hypoglycemia. INS-treated rats exhibited diminished LHA ORX gene expression and increased [ARH; LHA] or decreased [PVH] tissue ORX protein levels. 6-OHDA+INS animals showed a comparable decline in ORX transcripts, but attenuated augmentation of ARH and LHA ORX content and elevated PVH ORX levels. OT mRNA and protein were respectively decreased or unchanged during hypoglycemia, responses that were uninfluenced by hindbrain CA nerve cell destruction. These results illustrate divergent adjustments in glucoregulatory neurotransmitter gene expression and site-specific protein accumulation in the hypothalamus during hypoglycemia. Evidence that 6-OHDA pretreatment does not modify NPY or ORX transcriptional reactivity to hypoglycemia, but alters hypoglycemic patterns of NPY and ORX accretion implicates dorsomedial hindbrain CA neurons in regulation of translation/post-translational processing and site-specific availability of these neurotransmitters in the hypothalamus during hypoglycemia.

Topics: Animals; Blood Glucose; Blotting, Western; Catecholamines; Glucose; Hydroxydopamines; Hypoglycemia; Hypoglycemic Agents; Hypothalamus; Immunohistochemistry; Insulin; Intracellular Signaling Peptides and Proteins; Male; Neuropeptide Y; Neuropeptides; Neurotoxins; Neurotransmitter Agents; Orexins; Oxytocin; Polymerase Chain Reaction; Rats; Rats, Sprague-Dawley; Rhombencephalon; RNA, Messenger; Sympathectomy, Chemical

The 5q14.3 deletion syndrome is a rare chromosomal disorder characterized by moderate to severe intellectual disability, seizures and dysmorphic features. We report a 14-year-old boy with 5q14.3 deletion syndrome who carried a heterozygous deletion of the myocyte-specific enhancer factor 2c (MEF2C) gene. In addition to the typical neurodevelopmental features of 5q14.3 deletion syndrome, he showed recurrent hypoglycemia, appetite loss and hypothermia. Hormonal loading tests using insulin, arginine and growth hormone-releasing factor revealed that growth hormone was insufficiently released into serum in response to these stimuli, thus disclosing the hypothalamic dysfunction in the present case. To uncover the biological roles of MEF2C in the hypothalamus, we studied its expression in the postnatal mouse brain. Notably, neuropeptide Y (NPY)-positive interneurons in the hypothalamic arcuate nuclei highly expressed MEF2C. In contrast, the Rett syndrome-associated protein, Methyl-CpG binding Protein 2 (MECP2) was barely expressed in these neurons. MEF2C knockdown or overexpression experiments using Neuro2a cells revealed that MEF2C activated the endogenous transcription of NPY. Conversely, siRNA-mediated knockdown of MECP2 led to derepression of the Npy gene. These data support the concept that MEF2C and MECP2 share common molecular pathways regulating the homeostatic expression of NPY in the adult hypothalamus. We propose that individuals with 5q14.3 deletion syndrome may exhibit neuroendocrine phenotypes through the functional loss of MEF2C in the postnatal hypothalamus.

Topics: Abnormalities, Multiple; Adolescent; Animals; Chromosome Disorders; Chromosomes, Human, Pair 5; Female; Gene Deletion; Humans; Hypoglycemia; Hypothalamus; Hypothermia; Male; MEF2 Transcription Factors; Methyl-CpG-Binding Protein 2; Mice; Neuropeptide Y; Phenotype; Syndrome

The present study was undertaken to establish whether oxytocin (OT) is able to modify the NPY response to insulin-induced hypoglycemia in man. At 8:00 AM of 2 different days at least 1 week apart, 10 normal men were tested with insulin (0.15 IU/kg) and with the administration of OT (infused from time -15-60 min, at a constant rate of 2 mIU/ml) or placebo. Plasma NPY concentrations rose significantly during insulin tolerance test (ITT). Oxytocin treatment significantly reduced the NPY response to hypoglycemia. The finding demonstrates for the first time in humans that the systemic administration of OT exerts an inhibitory effect on the NPY rise caused by insulin-induced hypoglycemia.

Topics: Adult; Humans; Hypoglycemia; Insulin; Male; Neuropeptide Y; Oxytocin

The present study was undertaken in order to establish whether somatostatin (SRIH) is able to modify the neuropeptide Y (NPY) response to insulin-induced hypoglycemia during insulin tolerance test (ITT) in man. In addition, the possible involvement of opioid peptides in the mediation of hypoglycemia and/or SRIH action was investigated. Subjects were injected intravenously with 0.15IU/kg insulin alone (control test) or with SRIH (4.1μg/min/90min), naloxone (10mg in an iv bolus) or the combination of the two substances. Plasma NPY concentrations rose significantly during ITT. The NPY response was significantly reduced by the treatment with SRIH. The administration of naloxone did not modify NPY levels whereas when both SRIH and naloxone were given, NPY response to hypoglycemia did not differ from that observed in the control test. These data demonstrate that SRIH inhibits the NPY response to hypoglycemia. Naloxone-sensitive endogenous opiates do not seem to be involved in the control of hypoglycemia-induced NPY release. In contrast, since naloxone reversed the inhibiting effect of SRIH, an involvement of opioid peptides in the SRIH action may be supposed.

Topics: Adult; Blood Glucose; Hematocrit; Humans; Hypoglycemia; Insulin; Male; Naloxone; Narcotic Antagonists; Neuropeptide Y; Opioid Peptides; Somatostatin

The maintenance of appropriate glucose levels is necessary for survival. Within the brain, specialized neurons detect glucose fluctuations and alter their electrical activity. These glucose-sensing cells include hypothalamic arcuate nucleus neurons expressing neuropeptide Y (NPY) and lateral hypothalamic area (LHA) neurons expressing orexin/hypocretins (ORX) or melanin-concentrating hormone (MCH). Within the LHA, a population of NPY-expressing cells exists; however, their ability to monitor energy status is unknown. We investigated whether NPY neurons located in the LHA, a classic hunger center, detect and respond to fluctuations in glucose availability and compared these responses with those of known LHA glucose sensors expressing ORX or MCH. Using mice expressing green fluorescent protein under the control of NPY regulatory elements, we identified LHA NPY cells and explored their anatomical distribution, neurochemical and electrical properties, in vivo responses to fasting and insulin-induced hypoglycemia, and in situ electrical responses to extracellular glucose. We report that NPY, ORX, and MCH are expressed in nonoverlapping populations within the LHA. Subpopulations of LHA NPY neurons were activated in vivo by both a 6-h fast and insulin-induced hypoglycemia. Likewise, increased extracellular glucose suppressed the electrical activity of approximately 70% of LHA NPY neurons in situ, eliciting hyperpolarization and activating background K+ currents. Furthermore, we report that the glucose sensitivity of LHA NPY neurons is significantly different from neighboring ORX and MCH neurons. These data suggest that NPY-expressing cells in the LHA are a novel population of glucose-sensing neurons that represent a new player in the brain circuitry integrating information about glucose homeostasis.

Topics: Animals; Female; Food Deprivation; Glucose; Hypoglycemia; Hypothalamic Area, Lateral; Hypothalamic Hormones; Intracellular Signaling Peptides and Proteins; Male; Melanins; Mice; Mice, Transgenic; Neurons; Neuropeptide Y; Neuropeptides; Orexins; Pituitary Hormones

Many signals reflecting energy balance and stress are integrated at the hypothalamic orexigenic NPY neurons. To determine transcriptional changes of the NPY gene in response to stress, we followed the time course and compared the expression of heteronuclear (hn)- and messenger (m)RNA levels by in situ hybridization histochemistry and by real time PCR in mice following insulin-induced hypoglycemia and restraint. Hypoglycemia in fasted mice resulted in a rapid increase of NPY hnRNA that peaked at 1h, declined thereafter by 2-4h after insulin injection and run parallel to that of NPY mRNA. Throughout the time course examined, NPY expressing cells in the medial basal hypothalamus remained overwhelmingly localized to the arcuate nucleus. Following restraint NPY mRNA slightly increased, however hnRNA levels decreased up to 2h, suggesting increased stability of mature NPY mRNA. These results highlight rapid changes and differential regulation of NPY expression in response to metabolic and stress challenges.

Topics: Animals; Blood Glucose; Gene Expression Regulation; Hypoglycemia; Hypothalamus; Immunohistochemistry; In Situ Hybridization; Insulin; Male; Mice; Neuropeptide Y; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stress, Psychological; Time Factors; Transcriptional Activation; Up-Regulation

Arcuate neuropeptide Y (NPY)/agouti-related pepide (AgRP) neurones regulate energy homeostasis, and express the putative glucosensor, glucokinase (GCK). The present study performed multi-transcriptional profiling of these neurones to characterise NPY, AgRP and GCK gene expression during intermediate insulin-induced hypoglycaemia, and to determine whether these transcriptional responses acclimate to repeated insulin dosing. We also examined whether these neurones express insulin, glucocorticoid and oestrogen receptor gene transcripts, and whether the levels of these receptor mRNAs are modified by insulin-induced hypoglycaemia. Individual NPY-immunoreactive neurones were laser-microdissected from the caudal arcuate nucleus after single or serial dosing with neutral protamine Hagedorn insulin (NPH), and evaluated by quantitative real-time reverse transcriptase-polymerase chain reaction for the assessment of neurotransmitter and receptor gene expression. Mean NPY and AgRP mRNA in harvested NPY neurones was unchanged or augmented, respectively, by one NPH dose, although repeated NPH administration up-regulated NPY, whereas AgRP gene transcripts were down-regulated. NPH elicited divergent modifications in the ERalpha and ERbeta mRNA content of sampled neurones. ERalpha transcripts were amplified by both acute and chronic NPH-induced hypoglycaemia, whereas ERbeta gene expression was unaltered during a single bout, but suppressed during recurring hypoglycaemia. Glucocorticoid receptor (GR) mRNA levels were increased by a single insulin dose, but unaffected by serial NPH dosing. Insulin receptor-beta chain (InsRb) gene transcripts were insensitive to acute NPH-induced hypoglycaemia, but repeated NPH inhibited this gene transcript. Neither acute nor recurring hypoglycaemia modified GCK mRNA levels in caudal hypothalamic arcuate nucleus (ARH) NPY/AgRP neurones, but baseline GCK transcription was suppressed by the latter. This evidence for the habituation of hypoglycaemic patterns of InsRb, GR and ERbeta gene transcription to serial NPH dosing implies that such treatment may alter reactivity of caudal ARH NPY/AgRP neurones to receptor ligands, and supports the need to determine whether adaptive changes in neuronal sensitivity to insulin, corticosterone and/or oestrogen cause up- versus down-regulation of NPY and AgRP neurotransmission, respectively, by this caudal ARH subpopulation during chronic hypoglycaemia.

Topics: Agouti-Related Protein; Animals; Arcuate Nucleus of Hypothalamus; Base Sequence; DNA Primers; Estrogen Receptor alpha; Estrogen Receptor beta; Female; Hypoglycemia; Insulin, Isophane; Lasers; Neurons; Neuropeptide Y; Polymerase Chain Reaction; Rats; Rats, Sprague-Dawley

To assess mechanisms relating to food intake and glucosensing in fish, and their interaction with stress, we evaluated changes in the expression of orexigenic (NPY) and anorexigenic (POMC, CART and CRF) peptides in central glucosensing areas (hypothalamus and hindbrain) of rainbow trout subjected to normoglycaemic (control), hypoglycaemic (4 mg insulin kg(-1)) or hyperglycaemic (500 mg glucose kg(-1)) conditions for 6 h under normal stocking density (NSD; 10 kg fish mass m(-3)) or under stress conditions induced by high stocking density (HSD; 70 kg fish mass m(-3)). Hyperglycaemic NSD conditions resulted in decreased mRNA levels of NPY and increased levels of CART and POMC in the hypothalamus as well as increased mRNA levels of CART and CRF in the hindbrain compared with hypo- and normoglycaemic conditions. HSD conditions in normoglycaemic fish induced marked changes in the expression of all peptides assessed: mRNA levels of NPY and CRF increased and mRNA levels of POMC and CART decreased in the hypothalamus, whereas the expression of all four peptides (NPY, POMC, CART and CRF) decreased in the hindbrain. Furthermore, HSD conditions altered the response to changes in glycaemia of NPY and POMC expression in the hypothalamus and CART expression in the hypothalamus and the hindbrain. The results are discussed in the context of food intake regulation by glucosensor systems and their interaction with stress in fish.

Topics: Animals; Base Sequence; Blood Glucose; Corticotropin-Releasing Hormone; Crowding; DNA Primers; Eating; Fish Proteins; Gene Expression; Hyperglycemia; Hypoglycemia; Hypothalamus; Nerve Tissue Proteins; Neuropeptide Y; Neuropeptides; Oncorhynchus mykiss; Pro-Opiomelanocortin; Rhombencephalon; RNA, Messenger; Stress, Physiological

Expression of neuropeptide Y (Npy) heteronuclear (hn) RNA, an indicator of gene transcription, was significantly increased in the arcuate nucleus of rats 30min after insulin injection. Npy hnRNA levels were also increased significantly in response to hypoglycemia in rats in which the hypothalamus was deafferentated, although the absolute levels were significantly lower than in sham-operated rats. Direct effects of lowering glucose levels on Npy gene expression were also confirmed in hypothalamic organotypic cultures. Thus, Npy gene transcription in the arcuate nucleus increases rapidly in response to hypoglycemia, and both direct and indirect inputs are involved in the rapid upregulation.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Blood Glucose; Culture Media; Gene Expression; Gene Expression Regulation; Glucose; Hypoglycemia; Male; Neuropeptide Y; Organ Culture Techniques; Rats; Rats, Sprague-Dawley

Animals respond to hypoglycemia by eating and by stimulating gluconeogenesis. These responses to glucose deprivation are initiated by glucose-sensing neurons in the brain, but the neural circuits that control feeding behavior are not well established. Neurons in the arcuate region of the hypothalamus that express neuropeptide Y (NPY) and agouti-related protein (AgRP) have been implicated in mediating the feeding response to glucoprivation. We devised a method to selectively ablate these neurons in neonatal mice and then tested adult mice for their feeding responses to fasting, mild hypoglycemia, 2-deoxy-d-glucose and a ghrelin receptor agonist. Whereas the feeding response to the ghrelin receptor agonist was completely abrogated, the feeding response to glucoprivation was normal. The feeding response after a fast was attenuated when standard chow was available but normal with more palatable solid or liquid diet. We conclude that NPY/AgRP neurons are not necessary for generating or mediating the orexigenic response to glucose deficiency, but they are essential for the feeding response to ghrelin and refeeding on standard chow after a fast.

Topics: Agouti-Related Protein; Animals; Feeding Behavior; Female; Hypoglycemia; Insulin; Intercellular Signaling Peptides and Proteins; Male; Mice; Mice, Inbred C57BL; Neurons; Neuropeptide Y; Receptors, G-Protein-Coupled; Receptors, Ghrelin

The ovarian steroid hormone, estradiol, is one of several peripheral metabolic signal modulators that are integrated at the level of the arcuate nucleus of the hypothalamus (ARH), and is implicated in the control of ARH neuropeptides that maintain energy balance, including neuropeptide Y (NPY) and proopiomelanocortin (POMC). The present studies utilized quantitative real-time RT-PCR techniques to examine the hypothesis that estradiol regulates ARH NPY, POMC, and cocaine- and amphetamine-related transcript (CART) gene expression during acute insulin-induced hypoglycemia (IIH) and that adaptive modifications in transcriptional reactivity during recurring exposure are steroid dependent. ARH tissue was obtained by micropunch dissection from estradiol benzoate- and oil-implanted ovariectomized (OVX) rats that were treated by subcutaneous injection of one or four doses of the intermediate insulin formulation, Humulin NPH, over as many days, or vehicle alone. Our data show that in OVX plus estradiol benzoate and OVX plus oil groups, a single injection of insulin did not modify gene expression profiles, with the exception of acute hypoglycemic reduction of ARH NPY transcripts in the presence of estrogen. Prior exposure to daily hypoglycemia significantly diminished basal NPY and POMC mRNA levels in estradiol benzoate-, but not oil-implanted OVX rats, but elevated baseline CART transcripts in oil-treated animals. Recurring IIH enhanced ARH NPY gene expression relative to baseline, irrespective of the estradiol manipulation, but net tissue levels were greater in the absence of estrogen. In contrast, reexposure to hypoglycemia decreased POMC and CART gene transcription in estradiol benzoate- and oil-implanted OVX animals, respectively, relative to the single-dose groups. These studies show that estrogen modulates the impact of precedent exposure to IIH on basal and/or hypoglycemia-associated patterns of expression of ARH neuropeptide genes of characterized significance for energy homeostasis. The novel evidence for transcriptional acclimation of NPY, POMC, and CART to recurring IIH supports the possibility that adaptation of compensatory behavioral and physiological responses to acute versus chronic exposure to this metabolic stress may reflect neural regulatory mechanisms involving one or more neurotransmitters encoded by these genes.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Estradiol; Female; Gene Expression Regulation; Hypoglycemia; Insulin; Nerve Tissue Proteins; Neuropeptide Y; Ovariectomy; Pro-Opiomelanocortin; Rats; Rats, Sprague-Dawley; Transcription, Genetic

The mechanism of onset of hypoglycemia in patients with carnitine deficiency has yet to be determined. Using mice with systemic carnitine deficiency (JVS mice), we examined this mechanism, focusing on the weaning period (days 14-28 postpartum). For normal mice, the survival rate was 100%, and no hypoglycemia was observed at all. Gastric lactose began to decrease on day 17, and cellulose increased sharply in amount thereafter. For JVS mice, the survival rate was 77% on day 14 and 28% on day 28. From day 21 on, hypoglycemia was noted. Gastric lactose had disappeared almost completely by day 17, and cellulose was almost undetectable from days 14 to 28. Expression of orexin mRNA in the hypothalamus did not differ between JVS and normal mice on day 14, but was suppressed in JVS mice on days 21 and 28. When JVS mice were fed a carnitine-rich diet, suppression of expression of orexin mRNA in hypothalamus was eliminated, and on day 28 lactose and cellulose were detected in the stomach without hypoglycemia. In conclusion, the suppression of the expression of orexin in the hypothalamus during the weaning period may be involved in the marked anorexia in JVS mice, which eventually leads to death from hypoglycemia.

Topics: 3-Hydroxybutyric Acid; Animals; Behavior, Animal; Blood Glucose; Carnitine; Cellulose; Darkness; Female; Hypoglycemia; Hypothalamus; Intracellular Signaling Peptides and Proteins; Lactose; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Neuropeptide Y; Neuropeptides; Nucleic Acid Hybridization; Orexins; Pregnancy; Pro-Opiomelanocortin; Specific Pathogen-Free Organisms

Circulating glucose levels significantly affect vagal neural activity, which is important in the regulation of pancreatic functions. Little is known about the mechanisms involved. This study investigates the neural pathways responsible for hypoglycemia-induced vagal efferent signaling to the pancreas and identifies the neurotransmitters involved. Vagal pancreatic efferent nerve activities were recorded in anesthetized rats. Insulin-induced hypoglycemia, a decrease of blood glucose levels from 114 +/- 5 to 74 +/- 6 mg dl(-1), stimulated an increase in pancreatic efferent nerve firing from a basal rate of 1.1 +/- 0.3 to 19 +/- 3 impulses 30 s(-1). In contrast, vagal primary afferent neuronal discharges recorded in the nodose ganglia were unaltered by systemic hypoglycemia. Vagal afferent rootlet section plus splanchnicotomy had no effect on hypoglycemia-induced vagal efferent firing, suggesting a central site of action. Decerebration reduced the increase in nerve firing stimulated by hypoglycemia from 21 +/- 4 to 9.6 +/- 2 impulses 30 s(-1). Chemical ablation of the lateral hypothalamic area, but not the arcuate nucleus, inhibited pancreatic nerve firing evoked by hypoglycemia. Microinjection of the orexin-A receptor antagonist SB-334867 into the dorsal motor nucleus of the vagus (DMV) inhibited pancreatic nerve firing evoked by insulin-induced hypoglycemia by 56%. In contrast, injection of orexin-A (20 pmol) into the DMV elicited a 30-fold increase in pancreatic nerve firing. We concluded that systemic hypoglycemia stimulates pancreatic efferent nerve firing through a central mechanism. Full expression of pancreatic nerve activities during hypoglycemia requires both the forebrain and the brain stem. In addition to activating neurons in the brain stem, central neuroglucopenia activates subpopulations of neurons in the lateral hypothalamic area that contain orexin. The released orexin acts on DMV neurons to stimulate pancreatic efferent nerve activities and thus regulate pancreatic functions.

Topics: Action Potentials; Animals; Benzoxazoles; Blood Glucose; Brain Stem; Carrier Proteins; Dose-Response Relationship, Drug; Electric Stimulation; Electrophysiology; Excitatory Amino Acid Agonists; Glucose; Hypoglycemia; Hypothalamus; Immunohistochemistry; Insulin; Intracellular Signaling Peptides and Proteins; Kainic Acid; Male; Microinjections; Naphthyridines; Neural Pathways; Neurons, Efferent; Neuropeptide Y; Neuropeptides; Nodose Ganglion; Orexins; Pancreas; Rats; Rats, Sprague-Dawley; Serotonin; Splanchnic Nerves; Urea; Vagus Nerve

To investigate the role played by the orexigenic peptide, neuropeptide Y (NPY), in adaptive responses to insulin-induced hypoglycemia, we measured hypothalamic, feeding, and hormonal responses to this stimulus in both wild-type (Npy+/+) and NPY-deficient (Npy-/-) mice. After administration of insulin at a dose (60 mU ip) sufficient to cause moderate hypoglycemia (plasma glucose levels, 40 +/- 3 and 37 +/- 2 mg/dl for Npy+/+ and Npy-/- mice, respectively; P = not significant), 4-h food intake was increased 2.5-fold in Npy+/+ mice relative to saline-injected controls. By comparison, the increase of intake in Npy-/- mice was far smaller (45%) and did not achieve statistical significance (P = 0.08). Hyperphagic feeding in response to insulin-induced hypoglycemia was therefore markedly attenuated in mice lacking NPY, and a similar feeding deficit was detected in these animals after neuroglucopenia induced by 2-deoxyglucose (500 mg/kg ip). A role for NPY in glucoprivic feeding is further supported by our finding that Npy mRNA content (measured by real-time PCR) increased 2.4-fold in the hypothalamus of Npy+/+ mice by 7 h after insulin injection. Unlike the feeding deficits observed in mice lacking NPY, the effect of hypoglycemia to increase plasma glucagon and corticosterone levels was fully intact in these animals, as were both the nadir glucose value and time to recovery of euglycemia after insulin injection (P = not significant). We conclude that NPY signaling is required for hyperphagic feeding, but not neuroendocrine responses to moderate hypoglycemia.

Topics: Animals; Antimetabolites; Blood Glucose; Corticosterone; Deoxyglucose; Feeding Behavior; Female; Glucagon; Hyperphagia; Hypoglycemia; Hypothalamus; Male; Mice; Mice, Inbred Strains; Mice, Knockout; Neuropeptide Y; Signal Transduction

Rats that develop diet-induced obesity (DIO) on a 31% fat [high-energy (HE)] diet have defective sensing and responding to altered glucose levels compared with diet-resistant (DR) rats. Thus we postulated that they would also have defective counterregulatory responses (CRR) to insulin-induced hypoglycemia (IIH). Chow-fed selectively bred DIO and DR rats underwent three sequential 60-min bouts of IIH separated by 48 h. Glucose levels fell comparably, but DIO rats had 22-29% lower plasma epinephrine (Epi) levels during the first two bouts than DR rats. By the third trial, despite comparable Epi levels, DIO rats had lower 30-min glucose levels and rebounded less than DR rats 85 min after intravenous glucose. Although DIO rats gained more carcass and fat weight after 4 wk on an HE diet than DR rats, they were unaffected by prior IIH. Compared with controls, DR rats with prior IIH and HE diet had higher arcuate nucleus neuropeptide Y (50%) and proopiomelanocortin (POMC; 37%) mRNA and an inverse correlation (r = 0.85; P = 0.004) between POMC expression and body weight gain on the HE diet. These data suggest that DIO rats have a preexisting defect in their CRR to IIH but that IIH does not affect the expression of their hypothalamic neuropeptides or weight gain as it does in DR rats.

Topics: Adipose Tissue; Animals; Arcuate Nucleus of Hypothalamus; Blood Glucose; Body Composition; Catecholamines; Diet; Homeostasis; Hypoglycemia; Hypoglycemic Agents; Hypothalamus; In Situ Hybridization; Insulin; Lactic Acid; Male; Neuropeptide Y; Obesity; Organ Size; Pro-Opiomelanocortin; Rats; RNA, Messenger; Weight Gain

Topics: Adult; Diabetes Mellitus, Type 1; Diabetic Neuropathies; Female; Heart Rate; Humans; Hypoglycemia; Insulin; Male; Neuropeptide Y; Pancreatic Polypeptide; Respiratory Mechanics

Hypoglycemia reduces sympathoadrenal responses to subsequent hypoglycemic bouts by an unknown mechanism. To assess whether such hypoglycemia-associated autonomic failure is due to actual brain damage, male Sprague-Dawley rats underwent 1-h bouts of insulin-induced (5 U/kg i.v.) hypoglycemia (1.6-2.8 mmol/l) 1 or 3 times on alternate days. Rats remained alert and were rescued with intravenous glucose at 60-80 min. Plasma epinephrine and corticosterone responses were significantly reduced during the second and third bouts. Brains from these rats were processed by the terminal transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling (TUNEL) procedure as an index of apoptotic cell death at 24, 48, or 96 h after their first bout. At 48 h, but not 24 h, TUNEL+ cells were consistently seen only in the arcuate nucleus (arcuate hypothalamic nucleus [ARC]). Hypoglycemic rats had 188% more apoptotic ARC cells (1 bout 39+/-5; 3 bouts 37+/-4) than euglycemic controls (13+/-3;P = 0.001). In situ hybridization for neuropeptide Y (NPY) and proopiomelanocortin (POMC) mRNA was performed in sections of ARC containing maximal numbers of apoptotic cells as well as in other fresh frozen brains. After 1 bout, NPY (0.041+/-0.003) and POMC (0.119+/-0.022) mRNA were decreased, respectively, by 52 and 55% vs. controls (NPY 0.076+/-0.007; POMC 0.222+/-0.020; P = 0.01). NPY (0.029+/-0.002) but not POMC (0.093+/-0.013) fell 29% further after a third bout. NPY (r = -0.721; P = 0.001) and POMC (r = -0.756; P = 0.001) mRNA levels correlated negatively with the number of apoptotic ARC cells in the same sections. Thus, non-coma hypoglycemia produces apparent apoptotic cell death with reduced NPY and POMC expression selectively in the ARC. This may contribute to the reduced counterregulatory response following repeated bouts of hypoglycemia.

Topics: Animals; Apoptosis; Arcuate Nucleus of Hypothalamus; Brain; Hypoglycemia; Hypoglycemic Agents; In Situ Hybridization; In Situ Nick-End Labeling; Insulin; Male; Neuropeptide Y; Pro-Opiomelanocortin; Rats; Rats, Sprague-Dawley; RNA, Messenger

Maternal low protein malnutrition during gestation and lactation (LP) is an animal model frequently used for the investigation of long-term deleterious consequences of perinatal growth retardation. Both perinatal malnutrition and growth retardation at birth are risk factors for diabetic and cardiovascular disturbances in later life. The pathophysiologic mechanisms responsible are unknown. Hypothalamic nuclei are decisively involved in the central nervous regulation of food intake, body weight and metabolism. We investigated effects of a low protein diet (8% protein; control diet, 17% protein) during gestation and lactation in rat dams on the organization of hypothalamic regulators of body weight and metabolism in the offspring at weaning (d 20 of life). LP offspring had significantly lower body weight than control offspring (CO; P: < 0.001), associated with hypoglycemia and hypoinsulinemia (P: < 0. 005) on d 20 of life. This was accompanied by a greater relative volume of the ventromedial hypothalamic nucleus (P: < 0.01) and a greater numerical density of Nissl-stained neurons in this nucleus (P: < 0.01) as well as in the paraventricular hypothalamic nucleus (PVN; P: < 0.001). In contrast, no significant differences in neuronal densities were observed generally in the lateral hypothalamic area, arcuate hypothalamic nucleus (ARC), and dorsomedial hypothalamic nucleus between LP offspring and CO offspring. On the other hand, LP offspring displayed fewer neurons immunopositive for neuropeptide Y in the ARC (P: < 0.05), whereas in the PVN, lower neuronal densities of neurons immunopositive for galanin were found in LP offspring compared with CO offspring (P: < 0.001). On the contrary, in the PVN, no significant group difference in the numerical density of cholecystokinin-8S-positive neurons was present. A long-term effect of these specific hypothalamic alterations on body weight and metabolism in LP offspring during later life is suggested.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Body Weight; Female; Galanin; Hypoglycemia; Hypothalamus; Immunohistochemistry; Insulin; Lactation; Neurons; Neuropeptide Y; Paraventricular Hypothalamic Nucleus; Pregnancy; Pregnancy Complications; Protein Deficiency; Rats; Rats, Wistar; Ventromedial Hypothalamic Nucleus; Weaning

Hypoglycemia causes hyperphagia and weight gain, through unknown peripheral and central signals. We investigated the effect of hypoglycemia on NPY and leptin expression and the ability of leptin to inhibit hypoglycemia-induced hyperphagia. Acute hypoglycemia (60 U/kg SC insulin; n = 8) increased food intake (p < 0.01) compared with controls (n = 8). Insulin- and leptin-treated rats (300 microg/kg IP leptin; n = 8) had reduced hyperphagia (p < 0.05 vs. controls; p < 0.05 vs. insulin alone) and a 15% fall in NPY mRNA levels compared with controls (p < 0.01). Chronic hypoglycemia, (20-60 U/kg/day insulin; n = 8) increased food intake compared with vehicle-treated controls (p < 0.01). Leptin and insulin administration (300 microg/kg/day IP leptin; n = 8) reduced hyperphagia (p < 0.01 vs. controls, p < 0.05 vs. insulin alone), and NPY mRNA fell by 18% vs. controls (p < 0.01). We conclude that hypoglycemia-induced hyperphagia is not mediated by either a fall in leptin or an increase in hypothalamic NPY mRNA. Leptin can inhibit feeding in hyperphagic hypoglycemic rats, and this may partly be attributable to its inhibition of the NPY neurons.

Topics: Acute Disease; Analysis of Variance; Animals; Arcuate Nucleus of Hypothalamus; Blood Glucose; Body Weight; Chronic Disease; Eating; Hyperphagia; Hypoglycemia; Insulin; Leptin; Male; Models, Biological; Neuropeptide Y; Protein Biosynthesis; Proteins; Rats; Rats, Wistar; RNA, Messenger

The neuropeptides enkephalin (ENK), galanin (GAL) and neuropeptide Y (NPY) are abundantly expressed in the paraaortic body (PAB) and adrenal glands of the newborn rabbit. To examine whether these neuropeptides are affected by acute stress, we exposed neonatal rabbits to asphyxia, insulin-induced hypoglycemia, and reserpine. Asphyxia, caused by rebreathing for 60 min in an airtight box, reduced the content of catecholamines (CAs) in the adrenal glands and increased ENK-like immunoreactivity (-LI) in the PAB. Insulin-induced hypoglycemia reduced the content of CAs as well as ENK-LI in the adrenal glands. Reserpine caused a marked depletion of the CAs both in the PAB and in the adrenal glands. In contrast, reserpine did not cause any change in the contents of the neuropeptides in either organ. These data indicate that tissue levels of the neuropeptides GAL-LI and NPY-LI, coexisting with CA in the PAB and the adrenal glands, are not biochemically affected by asphyxia, hypoglycemia or reserpine, whereas tissue levels of ENK-LI are reduced by hypoglycemia and, to some extent, are increased by asphyxia. Furthermore, even the CAs in the PAB were unaffected by asphyxia and hypoglycemia. Also, while reserpine reduces CA content, peptide levels are unaffected.

Topics: Adrenal Glands; Animals; Asphyxia; Blood Glucose; Catecholamines; Enkephalins; Galanin; Hypoglycemia; Insulin; Neuropeptide Y; Neuropeptides; Para-Aortic Bodies; Rabbits; Reserpine; Stress, Physiological

The pathogenetic mechanisms behind insulin resistance in polycystic ovary syndrome (PCOS) are far from fully elucidated. Aberrant counterregulatory responses to hypoglycaemia have been reported in patients with insulin resistance, and recent reports suggest that plasma glucose may be regulated at lower levels in women with PCOS. In this study we investigated the complete hormonal counterregulatory response to hypoglycaemia in women with PCOS.. Prospective cross-sectional study.. Eight obese (BMI > or = 25) and 10 non-obese (BMI < 25) women with PCOS, diagnosed by means of ultrasonography and clinical signs of chronic anovulation. Eight obese and 9 non-obese controls.. Hypoglycaemia was induced by an intravenous bolus of soluble insulin (0.15 IU/kg body weight). The counterregulatory responses of cortisol, GH, catecholamines, glucagon, chromogranin A (CGA), and neuropeptide Y (NPY) were studied together with symptoms of hypoglycaemia.. The obese women with PCOS had a more pronounced truncal-abdominal body fat distribution (waist hip ratio, WHR) and were hyperinsulinaemic, compared with the obese controls. All the women exhibited blood glucose levels (< 2 mmol/l) well below the threshold for the hormonal counterregulatory response and for the appearance of clinical symptoms. The non-obese women with PCOS showed a greater increase in serum concentrations of GH than the lean controls. The obese women with PCOS exhibited blunted responses of noradrenaline and NPY, but similar increases of adrenaline and CGA, compared with the obese controls. They also showed a lower symptom score during hypoglycaemia. The response of noradrenaline to hypoglycaemia correlated inversely with fasting insulin levels in the women with PCOS. Among all the obese women (PCOS and controls pooled) basal levels of noradrenaline correlated inversely with the WHR.. All the women with PCOS, independent of BMI, body fat distribution and insulin levels, showed preserved counterregulatory responses to hypoglycaemia. The reduced plasma levels of noradrenaline and the lower perception of hypoglycaemic symptoms in the obese women with PCOS could both reflect a lower activation of the sympathetic nervous system. This aberration seems related to truncal-abdominal obesity and hyperinsulinaemia. The finding of an increased response of GH in the lean women with PCOS could support previous suggestions of an altered activity of the GH/IGF-I system in these women.

Topics: Adult; Body Mass Index; C-Peptide; Chromogranin A; Chromogranins; Cross-Sectional Studies; Female; Growth Hormone; Humans; Hypoglycemia; Insulin; Neuropeptide Y; Norepinephrine; Obesity; Polycystic Ovary Syndrome; Prospective Studies

Our previous studies show that neuropeptide Y (NPY) is involved in mediating sympathetic nerve stimulation-induced vasoconstriction. Insulin hypoglycemia is known to produce increased sympathetic output and elevated arterial pressure. The present study examined the role of NPY in the hypertensive response to insulin by examining the effects of insulin on NPY gene expression, tissue content and release. Subcutaneous injection of insulin produced an immediate increase in plasma NPY immunoreactivity (NPYir) and delayed increases in adrenal and neuronal NPY mRNA and adrenal NPYir in rats. These results suggest that NPY may play a role in insulin-induced hypertension.

Topics: Adrenal Medulla; Animals; Denervation; Gene Expression Regulation; Hypertension; Hypoglycemia; Hypoglycemic Agents; Insulin; Male; Neuropeptide Y; Nicotine; Nicotinic Agonists; Rats; Rats, Sprague-Dawley; Stress, Physiological; Superior Cervical Ganglion

This study examined whether or not changes in plasma concentrations of motilin and other gastrointestinal hormones known to affect gastric motility are associated with the accelerated gastric emptying seen during hypoglycaemia. While studying gastric emptying by scintigraphy in eight healthy subjects, the plasma concentrations of glucagon, adrenaline, motilin, gastrin, neuropeptide Y and somatostatin were measured during normoglycaemia and hypoglycaemia with simultaneous infusion of either atropine or saline. Blood glucose concentrations were checked by an insulin-glucose clamp. The plasma levels of glucagon and adrenaline increased markedly during both hypoglycaemic examinations compared with normoglycaemia. Neither motilin nor any of the other hormones displayed considerable changes during hypoglycaemia with and without atropine compared with normoglycaemia. No further information about the mechanisms behind the accelerated gastric emptying rate during hypoglycaemia was obtained by analysing motilin and the other gastrointestinal hormones.

Topics: Adult; Blood Glucose; Epinephrine; Gastric Emptying; Gastrins; Glucagon; Glucose Clamp Technique; Humans; Hypoglycemia; Male; Motilin; Neuropeptide Y; Neuropeptides; Somatostatin

The changes in plasma gastrin-releasing peptide (GRP), arginine vasopressin (AVP), neuropeptide Y (NPY), corticotropin releasing hormone (CRH), galanin, ACTH, cortisol, delta sleep-inducing peptide (DSIP), adrenaline, noradrenaline and pancreatic polypeptide (PP) were measured after 5 and 15 minutes of acute insulin-induced moderate hypoglycaemia (2.0 mmol/l) in 10 patients with Type 1 diabetes mellitus with no autonomic neuropathy and in 10 healthy subjects. Plasma catecholamine and PP levels rose in both groups in response to hypoglycemia and the secretory response of ACTH was lower in the diabetic subjects (p < 0.01). GRP concentrations increased during hypoglycaemia (p < 0.01) while a reduction in AVP occurred at the start of hypoglycaemia (p < 0.001). The plasma AVP concentrations were higher in the diabetic group compared with those in the normal group (p < 0.05). The NPY concentrations were higher in the normal subjects (p < 0.05) but no change in the mean level occurred in either group during hypoglycaemia. No group differences or changes in mean plasma concentrations were found for galanin, DSIP and CRH. These observations support the view that regulatory peptides, if involved in glucose homeostasis, may rather have a modulatory effect than a direct action in restoring normoglycaemia.

Topics: Adrenocorticotropic Hormone; Adult; Analysis of Variance; Arginine Vasopressin; Blood Glucose; Diabetes Mellitus, Type 1; Epinephrine; Galanin; Gastrin-Releasing Peptide; Hormones; Humans; Hydrocortisone; Hypoglycemia; Neuropeptide Y; Norepinephrine; Pancreatic Polypeptide; Peptides; Reference Values; Time Factors

An immunoneutralization technique with specific antibodies was used to explore the role of endogenous neuropeptide Y (NPY) in the adrenocorticotropic hormone (ACTH) release after hypoglycemic stress in the dog. Dogs received injections of rabbit antihuman NPY gamma-globulin (anti-NPY) or normal gamma-globulin (NGG) into the third cerebral ventricle, which was followed by i.v. injection of insulin. Hypoglycemia of a 40% fall in systemic glucose levels occurred in anti-NPY-treated dogs as well as NGG-treated animals. An intraventricular administration of anti-NPY significantly inhibited the ACTH and cortisol release to hypoglycemia, but had no effect on the pancreatic polypeptide (PP) response. These findings suggest involvement by endogenous NPY in the ACTH secretion induced by hypoglycemia.

Topics: Adrenocorticotropic Hormone; Animals; Brain; Dogs; Hydrocortisone; Hypoglycemia; Immune Sera; Injections, Intraventricular; Insulin; Neuropeptide Y

Secretion of the adrenal medulla was stimulated in nine cats by insulin-induced hypoglycemia. Levels of catecholamines (mol. wt 153-183), neuropeptide Y (mol. wt 4254) and chromogranin A (mol. wt 48,000) were measured in concurrently collected samples of adrenolumbar venous blood and thoracic duct lymph for up to 4 h following insulin administration. Insulin-induced hypoglycemia elicited an increase in the secretion of catecholamines, which reached peak levels in the adrenolumbar venous plasma at 1.5-2 h and in the lymph at 2.5 h. Although catecholamines were the most numerous measured molecules in the lymph, levels of norepinephrine and epinephrine were 75-250-fold less than those found in the adrenolumbar venous plasma. Neuropeptide Y in the adrenolumbar venous plasma reached peak levels between 1 and 1.5 h; at this time approximately 20% of the peak venous amount was detected in the lymph. Chromogranin A was found in approximately equal amounts in both plasma and lymph; the peak level in the plasma occurred at 1.5-2 h, while that in the lymph was reached at 2-3 h. We suggest that the size of a molecule influences the route it takes following exocytosis from the chromaffin vesicle. Smaller molecules such as catecholamines may pass directly into the circulation, while larger molecules such as chromogranin A may be temporarily sequestered in the interstitial space before passing into the lymph, and hence into the circulation.

Topics: Adrenal Medulla; Animals; Catecholamines; Cats; Chromogranin A; Chromogranins; Hypoglycemia; Nerve Tissue Proteins; Neuropeptide Y; Thoracic Duct

Previous studies have demonstrated that plasma Neuropeptide Y-like immunoreactivity (NPY-LI) increases after activation of sympathetic nerves. To test the hypothesis that the adrenal medulla may also be a significant source of circulating plasma NPY-LI and to determine if NPY is co-released with adrenal catecholamines, we have measured the peripheral venous concentrations of NPY-LI, adrenaline and noradrenaline in six patients, before and after induction of hypoglycaemia as part of pituitary function tests that also tested gonadotrophin and thyroid stimulating hormone release. The plasma adrenaline concentration was increased approximately 15 times (p less than 0.05) relative to baseline at 30 mins and remained elevated for the 90 minutes of the study. The plasma concentration of both noradrenaline and NPY-LI remained unchanged. These results failed to demonstrate an increase in the amount of NPY-LI released into the plasma during stimulation of the adrenal medulla with hypoglycaemic stress in man. They do not support significant co-release of NPY with adrenaline from the adrenal medulla in man, nor a physiological role for NPY as an adrenal hormone in human subjects in this situation.

Topics: Adult; Aged; Female; Humans; Hypoglycemia; Male; Middle Aged; Neuropeptide Y

Peptide YY (PYY), neuropeptide Y, and pancreatic polypeptide (PP) are the third family of structurally related peptides to be extracted from the brain-gut axis. In the present study we have validated a sensitive and specific assay for PYY and used it to measure the distribution of PYY in the canine gastrointestinal tract. In addition we have compared the PYY and PP responses to both an intragastric meal and to insulin hypoglycemia. The highest concentration of PYY was present in the canine ileum (1610 +/- 123 ng/g) and colon (1607 +/- 194 ng/g). Significant (p less than 0.05) increases in both circulating PP and PYY concentrations were observed in response to the meal (mean delta PP = 180 +/- 43 pg/ml; mean delta PYY = 205 +/- 61 pg/ml). In contrast, only serum PP concentrations (mean delta PP = 294 +/- 36 pg/ml) increased significantly (p less than 0.01) in response to insulin hypoglycemia, demonstrating differences in the sensitivity of the PP and PYY cells to vagal stimulation. The present demonstration of PYY release in response to a physiological stimulus fulfills one of the prerequisites for hormonal status.

Topics: Amino Acid Sequence; Animals; Chromatography; Colon; Digestive System; Dogs; Eating; Gastrointestinal Hormones; Hypoglycemia; Ileum; Nerve Tissue Proteins; Neuropeptide Y; Pancreatic Polypeptide; Peptide YY; Peptides; Radioimmunoassay