piperidines and 4-fluorobenzyltrozamicol

Islet cell adaptation to insulin resistance in type 2 diabetes mellitus may be due in part to increased stimulation of beta cells by the autonomic nervous system. The parasympathetic neurotransmitter acetylcholine (ACh) mediates insulin release via M3 muscarinic receptors on islet beta cells. The vesicular ACh transporter (VAChT) receptor correlates with cholinergic activity in vivo. The positron emission tomography (PET) radiotracer (+)-4-[18F]fluorobenzyltrozamicol ([18F]FBT) binds to the VAChT receptor on presynaptic cholinergic neurons and can be quantified by PET. In this study, we utilize [18F]FBT PET to demonstrate pancreatic cholinergic activity before and after dextrose infusion in nonhuman primates with normal (NGT) and impaired (IGT) glucose tolerance.. Seven adult female vervet (Chlorocebus aethiops) monkeys were maintained on an atherogenic Western diet. They were divided into two groups: four with NGT and three with IGT. Each subject underwent [18F]FBT PET twice: first, a baseline PET under fasting conditions; and second, PET under fasting conditions but after intravenous infusion of dextrose solution. Quantitative analysis of pancreatic uptake at 60 min post-injection was performed.. There was no difference in pancreatic uptake of [18F]FBT on baseline scans between the two groups. Pancreatic uptake of [18F]FBT increased in every subject after dextrose infusion (P = 0.03). On post-dextrose PET scans, pancreatic uptake of [18F]FBT was significantly higher in IGT subjects compared with NGT subjects (P = 0.03). The post-dextrose to pre-dextrose uptake ratios were higher in IGT subjects (P = 0.08).. Acute increases in pancreatic cholinergic activity in vivo were detected in the pancreata of nonhuman primates with NGT and IGT after intravenous dextrose infusion on [18F]FBT PET. In subjects with IGT, this activity was significantly higher, suggesting increased autonomic nervous system stimulation of the pancreatic islets in insulin-resistant subjects.

Topics: Animals; Blood Glucose; Chlorocebus aethiops; Female; Fluorine Radioisotopes; Fluorobenzenes; Glucose Intolerance; Insulin; Piperidines; Positron-Emission Tomography; Primates; Reference Values; Triglycerides

Pancreatic neuronal changes associated with beta cell loss in type 1 diabetes mellitus are complex, involving, in part, parasympathetic mechanisms to compensate for preclinical hyperglycemia. The parasympathetic neurotransmitter acetylcholine (ACh) mediates insulin release via M3 muscarinic receptors on islet beta cells. The vesicular ACh transporter (VAChT) receptor has been shown to be a useful marker of cholinergic activity in vivo. The positron emission tomography (PET) radiotracer (+)-4-[(18)F]fluorobenzyltrozamicol ([(18)F]FBT) binds to the VAChT receptor on presynaptic cholinergic neurons and can be quantified by PET. The compound 4-diphenylacetoxy-N-methylpiperidine (4-DAMP), available in a tritiated form, binds to M3 muscarinic receptors on beta cells and is a potential target for assessing pancreatic beta cell mass. In this study, we investigate the feasibility of dual radiotracer analysis in identifying neurofunctional changes that may signify type 1 diabetes mellitus in its early preclinical state.. Ex vivo determinations of pancreatic uptake were performed in prediabetic nonobese diabetic mice and controls after intravenous injection of [(18)F]FBT or 4-[(3)H]DAMP. Beta cell loss in prediabetic mice was confirmed using immunohistochemical methods.. [(18)F]FBT uptake was significantly higher in prediabetic pancreata than controls: 3.22 +/- 0.81 and 2.51 +/- 1.04, respectively (P < 0.03). 4-[(3)H]DAMP uptake was significantly lower in prediabetic pancreata than controls: 0.612 +/- 0.161 and 0.968 +/- 0.364, respectively (P = 0.01).. These data suggest that a combination of radiotracer imaging agents that bind to neuronal elements intimately involved in insulin production may be an effective method of evaluating changes associated with early beta cell loss using PET.

Topics: Animals; Fluorine Radioisotopes; Fluorobenzenes; Insulin-Secreting Cells; Mice; Pancreas; Parasympatholytics; Piperidines; Prediabetic State; Radiography; Tritium

Vesamicol derivatives are promising candidates as ligands for the vesicular acetylcholine transporter (VAChT) to enable in vivo imaging of cholinergic deficiencies if applied as positron emission tomography radiotracers. So far, optimization of the binding affinity of vesamicol-type ligands was hampered by the lack of respective quantitative structure-activity relationships. We developed the first quantitative model to predict, from molecular structure, the binding affinity of vesamicol-type ligands toward VAChT employing comparative molecular field analysis (CoMFA) for a set of 37 ligands, covering three different structural types (4-phenylpiperidine, spiro, and tropan derivatives of vesamicol). The prediction capability was assessed by leave-one-out cross-validation (LOO) and through leaving out and predicting 50% of the compounds selected such that both the training and the prediction sets cover almost the whole range of experimental data. The statistics indicate a significant prediction power of the models ( q (2) (LOO) = 0.66, q (2) (50% out) = 0.59-0.74). The discussion includes detailed analyses of CoMFA regions critical for ligand-VAChT binding, identifying structural implications for high binding affinity.

Topics: Binding Sites; Fluorobenzenes; Iodobenzenes; Ligands; Models, Molecular; Molecular Structure; Piperidines; Quantitative Structure-Activity Relationship; Reproducibility of Results; Static Electricity; Stereoisomerism; Vesicular Acetylcholine Transport Proteins

The pancreas is one of the most heavily innervated peripheral organs in the body. Parasympathetic and sympathetic neurons terminate in the pancreas and provide tight control of endocrine and exocrine functions. The aim of this study was to determine whether the pancreas can be imaged with a radioligand that binds to specific neuroreceptors. Using fluorine-18 4-fluorobenzyltrozamicol (FBT), which binds to the presynaptic vesicular acetylcholine transporter, positron emission tomography scans were performed in four adult mice, two adult rhesus monkeys, and one adult human. In these mammals, the pancreas is intensely FBT avid, with uptake greater than in any other organ at 30, 60, and 90 min. The maximum standardized uptake value (SUV) ratios of pancreas to liver, for example, ranged from 1.4 to 1.7 in rhesus monkeys (mean 1.6; median 1.7) and from 1.9 to 4.7 (mean 3.24; median 3.02) in mice. The maximum SUV ratio of pancreas to liver in the human was 1.8. These data suggest that neuroreceptor imaging of the pancreas in vivo is feasible in animal models and humans. This imaging could allow researchers to interrogate functions under control of the autonomic nervous system in the pancreas, with applications possible in transplanted and native pancreata. Also, as beta cell function is intimately related to parasympathetic cholinergic input, FBT activity in the pancreas may correlate with insulin-producing beta cell mass. This could ultimately provide a method of in vivo imaging in animal models and humans for diabetes research.

Topics: Animals; Autonomic Nervous System; Female; Fluorobenzenes; Humans; Macaca mulatta; Male; Membrane Transport Proteins; Mice; Mice, Inbred BALB C; Neurons; Pancreas; Piperidines; Radiopharmaceuticals; Reproducibility of Results; Sensitivity and Specificity; Sensory Receptor Cells; Species Specificity; Tissue Distribution; Tomography, Emission-Computed; Vesicular Acetylcholine Transport Proteins; Vesicular Transport Proteins

In the present study, the radiotracer [(18)F] (+)-4-fluorobenzyltrozamicol ((+)-[(18)F]FBT) and positron emission tomography (PET) were used to examine the vesicular acetylcholine transporter and determine if presynaptic cholinergic activity was altered with age in 23 rhesus monkeys that varied in age from 10 to 37 years. Binding of (+)-[(18)F]FBT in the basal ganglia was reduced significantly with increasing age of the monkeys. However, there were individual differences noted in that some middle-aged and aged monkeys demonstrated levels of (+)-[(18)F]FBT binding that were comparable to the binding measured in adult monkeys. These data indicate that presynaptic cholinergic function may decrease with age, but that there may be a differential susceptibility of the cholinergic system to the aging process in different individuals.

Topics: Acetylcholine; Aging; Animals; Basal Ganglia; Carrier Proteins; Cerebellum; Fluorine Radioisotopes; Fluorobenzenes; Macaca mulatta; Male; Membrane Transport Proteins; Neuromuscular Depolarizing Agents; Piperidines; Presynaptic Terminals; Tomography, Emission-Computed; Vesicular Acetylcholine Transport Proteins; Vesicular Transport Proteins

Positron emission tomography (PET) imaging of spinal cord in monkeys with a cholinergic tracer demonstrates increased spinal cholinergic activity in response to an analgesic dose of morphine, and this PET result correlates with measurement of acetylcholine spillover into spinal cord extracellular space induced by morphine, as measured by microdialysis. Previous studies in rats, mice, and sheep demonstrate activation of spinal cholinergic neurons by systemic opioid administration, and participation of this cholinergic activity in opioid-induced analgesia. Testing the relevance of this observation in humans has been limited to measurement of acetylcholine spillover into lumbar cerebrospinal fluid. The purpose of this study was to apply a recently developed method to image spinal cholinergic terminals non-invasively via PET and to test the hypothesis that the tracer utilized would reflect changes in local cholinergic activity. Following Animal Care and Use Committee approval, seven adult male rhesus monkeys were anesthetized on three separate occasions. On two of the occasions PET scans were performed using [(18)F] (+)-4-fluorobenzyltrozamicol ([(18)F]FBT), which selectively binds to the vesicular acetylcholine (ACh) transporter in the presynaptic cholinergic terminals. PET scans were preceded by injection of either saline or an analgesic dose of IV morphine (10 mg/kg). On the third occasion, microdialysis catheters were inserted in the spinal cord dorsal horn and acetylcholine concentrations in dialysates determined before and after IV morphine injection. Morphine increased cholinergic activity in the spinal cord, as determined by blood flow corrected distribution volume of [(18)F]FBT in the cervical cord compared to the cerebellum. Morphine also increased acetylcholine concentrations in microdialysates from the cervical cord dorsal horn. The one animal which did not show increased spinal cholinergic activity by PET from this dose of morphine also did not show increased acetylcholine from this morphine dose in the microdialysis experiment. These data confirm the ability to use PET to image spinal cholinergic terminals in the monkey spinal cord and suggest that acute changes in cholinergic activity can be imaged with this non-invasive technique. Following preclinical screening, PET scanning with [(18)F]FBT may be useful to investigate mechanisms of analgesic action in normal humans and in those with pain.

Topics: Acetylcholine; Analgesics, Opioid; Animals; Cholinergic Fibers; Fluorine Radioisotopes; Fluorobenzenes; Macaca mulatta; Male; Microdialysis; Morphine; Piperidines; Regional Blood Flow; Spinal Cord; Tomography, Emission-Computed

[18F](+)-4-fluorobenzyltrozamicol (FBT), which selectively binds to the vesicular acetylcholine transporter in the presynaptic cholinergic neuron, has previously been shown to be a useful ligand for the study of cholinergic terminal density in the basal ganglia with PET. The goal of this study was to assess the test-retest variability of [18F]FBT and PET measurements under baseline conditions in the basal ganglia.. After approval from the Animal Care and Use Committee, 6 rhesus monkeys underwent a series of 2 [18F]FBT PET scans (time between scans, 32-301 d) under isoflurane anesthesia. Each scan was initiated on the bolus injection of the radiotracer and consisted of 26 frames acquired during 180 min. Arterial blood samples were collected over the course of each scan to determine the metabolite-corrected arterial input function. Tissue time-activity curves were obtained from the scan data by drawing regions of interest over the basal ganglia and cerebellum. The distribution volume ratio for the basal ganglia was then determined for each scan by taking the ratio of the basal ganglia (specific binding) to cerebellum (nonspecific binding) distribution volume. Distribution volumes were derived using the Logan graphic analysis technique as well as a standard 3-compartment model. Additionally, the radioactivity concentration ratio was calculated as the ratio of the average [18F]FBT concentration in the basal ganglia to that in the cerebellum during the last half of the study (85-170 min). The constant K1, determined using the standard 3-compartment model, was used as an index of blood flow changes between studies.. For all subjects, the test-retest variability was less than 15% for the distribution volume ratio and 12% for the radioactivity concentration ratio. Good agreement was found between the distribution volume ratio calculated using the graphic technique and the standard 3-compartment model. Using K1 as an index, the variability in blood flow seen in both the basal ganglia and the cerebellum was significantly reduced in their ratio.. These results show the reproducibility of [18F]FBT and PET measurements in the basal ganglia.

Topics: Animals; Basal Ganglia; Brain; Cerebellum; Cholinergic Fibers; Fluorine Radioisotopes; Fluorobenzenes; Macaca mulatta; Male; Nerve Endings; Piperidines; Radiopharmaceuticals; Reproducibility of Results; Tomography, Emission-Computed

The goal of the present set of studies was to characterize the in vitro binding properties and in vivo tissue kinetics for the vesicular acetylcholine transporter (VAcChT) radiotracer, [18F](+)-4-fluorobenzyltrozamicol ([18F](+)-FBT). In vitro binding studies were conducted in order to determine the affinity of the (+)- and (-)-stereoisomers of FBT for the VAcChT as well as sigma (sigma 1 and sigma 2) receptors. (+)-FBT was found to have a high affinity (Ki = 0.22 nM) for the VAcChT and lower affinities for sigma 1 (21.6 nM) and sigma 2 (35.9 nM) receptors, whereas (-)-FBT had similar affinities for the VAcChT and sigma 1 receptors (approximately 20 nM) and a lower affinity for sigma 2 (110 nM) receptors. PET imaging studies were conducted in rhesus monkeys (n = 3) with [18F](+)-FBT. [18F](+)-FBT was found to have a high accumulation and slow rate of washout from the basal ganglia, which is consistent with the labeling of cholinergic interneurons in this brain region. [18F](+)-FBT also displayed reversible binding kinetics during the 3 h time course of PET and produced radiolabeled metabolites that did not cross the blood-brain barrier. The results from the current in vitro and in vivo studies indicate that [18F](+)-FBT is a promising ligand for studying cholinergic terminal density, with PET, via the VAcChT.

Topics: Animals; Basal Ganglia; Brain; Carrier Proteins; Cerebellum; Fluorine Radioisotopes; Fluorobenzenes; Frontal Lobe; Macaca mulatta; Magnetic Resonance Imaging; Male; Membrane Transport Proteins; Neuromuscular Depolarizing Agents; Occipital Lobe; Organ Specificity; Piperidines; Radioligand Assay; Rats; Rats, Sprague-Dawley; Receptors, sigma; Stereoisomerism; Synaptic Vesicles; Temporal Lobe; Tomography, Emission-Computed; Vesicular Acetylcholine Transport Proteins; Vesicular Transport Proteins

Current methods of radioligand synthesis for the vesamicol receptor either utilize bioactive unlabeled precursors and/or generate unlabelled bioactive by-products. The presence of these compounds in the radiotracer preparation increases the risk of adverse pharmacological reactions in animals. To eliminate the risk of such reactions, we have synthesized the novel radioligand [18F]FBT from two inactive precursors, [18F]fluorobenzyl iodide and trozamicol. (+)- and (-)-[18F]FBT were synthesized in 57-85% yield, from [18F]fluorbenzyl iodide and (+)- and (-)-trozamicol, respectively. In the rat brain, comparable levels of (-)-[18F]FBT were found in the striatum, cortex, cerebellum and hippocampus during a 3 h period. In contrast, the striatum:cerebellum, cortex:cerebellum and hippocampus:cerebellum ratios for (+)-[18F]FBT increased from unity at 5 min post-injection to 2.8, 1.4 and 1.7, respectively, at 3 h. Therefore, (+)-[18F]FBT may be potentially useful for mapping vesamicol receptor density in vivo.

Topics: Animals; Brain; Cerebellum; Cerebral Cortex; Corpus Striatum; Fluorine Radioisotopes; Fluorobenzenes; Haloperidol; Hippocampus; Isotope Labeling; Male; Piperidines; Rats; Receptors, Cholinergic; Time Factors