preproenkephalin and Hypoglycemia

Glucoprivation or hypoglycemia induces a range of counterregulatory responses, including glucose mobilization, reduced glucose utilization, and de novo glucose synthesis. These responses are mediated in part by the sympathetic nervous system. The aim of this study was to determine the chemical codes of sympathetic preganglionic neurons (SPN) activated by glucoprivation, induced by 2-deoxy-D-glucose (2DG). SPN controlling the adrenal glands and celiac ganglia, which ultimately can innervate the liver and pancreas, were targeted together with the superior cervical ganglia (control). 23.9% ± 1.3% of SPN in the T4-T11 region contained c-Fos immunoreactivity following 2DG; 70.3% ± 1.8% of SPN innervating the adrenal glands and 37.4% ± 3% of SPN innervating celiac ganglia were activated. 14.8% ± 3.5% of SPN (C8-T3) innervating superior cervical ganglia were activated. In the C8-T3 region 55% ± 10% of SPN activated contained PPCART, with only 12% ± 3% expressing PPE mRNA, whereas, in the T4-T11 region, 78% ± 4% contained PPE, with only 6.0% ± 0.6% expressing PPCART mRNA. Thus CART is not involved in glucose mobilization. Two chemically distinct populations of SPN (PPE⁺ 57.4% ± 5%, PPE⁻ ∼40%) were identified to regulate adrenaline release in response to glucoprivation. Multiple chemically distinct SPN populations innervating a specific target could suggest their graded recruitment. The two distinct populations of SPN (PPE⁺ 67.6% ± 9%, PPE⁻ ∼30%) projecting to celiac ganglia activated by glucoprivation could direct pancreatic and hepatic or other counterregulatory responses. Nearly all SPN that expressed PPE mRNA and projected to the adrenal glands or celiac ganglia were activated, suggesting a role for the inhibitory peptide enkephalin in responses evoked by glucoprivation.

Topics: Adrenal Glands; Animals; Blood Glucose; Cholera Toxin; Choline O-Acetyltransferase; Chromaffin Cells; Deoxyglucose; Enkephalins; Epinephrine; Fluorescent Dyes; Ganglia, Sympathetic; Gene Expression Regulation; Hypoglycemia; Male; Nerve Tissue Proteins; Neurons; Phenylethanolamine N-Methyltransferase; Protein Precursors; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Spinal Cord

To further evaluate whether transsynaptic mechanisms account for stress-induced changes in adrenomedullary preproenkephalin mRNA (ppEnk mRNA), neonatal rats were made hypoglycemic at a time when synapses are non-functional (less than 10 days postnatal age). While ppEnk mRNA in medullae from adult rats increased as much as 60-fold in this paradigm (insulin 10 U/kg), ppEnk mRNA levels in the newborn increased only 1.6-fold (insulin 20 U/kg). To evaluate whether postsynaptic cholinergic pathways of the neonatal adrenal medulla were functional, we treated 5-day-old pups with cholinergic agonists (nicotine [1 mg/kg, s.c., q 12 h] + carbachol [1.7 mumol/kg, s.c., q 12 h x 4 days]). Combined cholinergic agonist treatment augmented enkephalin prohormone and peptide levels up to 3-fold (P less than 0.05). To determine whether the blunted response to hypoglycemia in the newborn resulted from a deficiency in functional transsynaptic activity, synapses were matured using thyroid hormone pretreatment (postnatal days 2 and 3) before hypoglycemic stress. Hypoglycemia now caused a 40-fold increase in adrenomedullary ppEnk mRNA levels only in the T3/insulin treated group. To exclude other secondary effects of hypoglycemia (eg. hormonal, or insulin treatment-dependent), intracellular glycopenia was produced in the presence of secondary hyperglycemia by injecting adult rats or pups with 2-deoxyglucose (500 mg/kg). Similar to the insulin-hypoglycemia group, a large increase in adrenomedullary ppEnk mRNA resulted in the adult but not in the 5-day-old neonatal adrenal medullae. We conclude that enkephalin biosynthesis, like co-stored catecholamines, is induced by a transsynaptic process.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adrenal Medulla; Animals; Animals, Newborn; Carbachol; Corpus Striatum; Deoxyglucose; Enkephalins; Gene Expression Regulation; Hypoglycemia; Insulin; Male; Nicotine; Protein Precursors; Rats; Rats, Inbred Strains; RNA, Messenger; Triiodothyronine

The significance of the 5' heterogeneity of the transmitter gene ppEnk was evaluated by comparing start site usage (E1-E4) between 12 tissues from untreated rats, using primer extension analysis. In the basal state, we found that E3- and E4-initiated transcripts accounted for 80% of the total striatal RNA present compared with a preferential usage of the E2 start site in all other tissues. To determine whether this selective expression could be modified by biologically relevant pathways, rats were made hypoglycemic. After insulin shock, only E3 + E4-initiated transcripts increased (16-fold at 1 day) in the adrenal medulla but were unaffected in the striatum. As the effects of insulin shock on the adrenal medulla are mediated by cholinergic pathways and the striatum also receives cholinergic inputs, we also compared the effects of cholinergic drug treatments on start site usage in these two tissues. Rats were treated with cholinergic agonists (nicotine + oxotremorine) which induced adrenomedullary E2 and E3 + E4 transcripts (5- and 80-fold, respectively). This effect peaked at 2 days. In contrast, in the same animals, striatal ppEnk RNA (E3 + E4) increased only 10-15-fold after drug treatment. Hence it appears that biologically relevant whole animal stimuli (insulin shock or cholinergic agents) activate biochemical pathways, which affect start site usage in a tissue-specific fashion. Selective RNA start site usage suggests a biological significance, which may be important in the widespread tissue expression of this gene.

Topics: Adrenal Medulla; Animals; Base Sequence; Corpus Striatum; Enkephalins; Gene Expression; Hypoglycemia; Male; Molecular Sequence Data; Nicotine; Oxotremorine; Protein Precursors; Rats; Rats, Inbred Strains; RNA, Messenger; Tissue Distribution; Transcription, Genetic