Sex differences in [123I]beta-CIT SPECT measures of dopamine and serotonin transporter availability in healthy smokers and nonsmokers.

Nicotine and other constituents of tobacco smoke elevate dopamine (DA) and serotonin (5-HT) levels in brain and may cause homeostatic adaptations in DA and 5-HT transporters. Since sex steroids alter DA and 5-HT transporter expression, the effects of smoking on DA and 5-HT transporter availability may differ between sexes. In the present study, DA and 5-HT transporter availabilities were quantitated using single photon emission computed tomography (SPECT) imaging approximately 22 h after bolus administration of [123I]beta-CIT, an analog of cocaine which labels DA and 5-HT transporters. Forty-two subjects including 21 pairs of age-, race-, and gender-matched healthy smokers and nonsmokers (12 female and 9 male pairs) were imaged. Regional uptake was assessed by the outcome measures, V3", which is the ratio of specific (i.e., ROI-cerebellar activity) to nondisplaceable (cerebellar) activity, and V3, the ratio of specific to free plasma parent. Overall, striatal and diencephalic [123I]beta-CIT uptake was not altered by smoking, whereas brainstem [123I]beta-CIT uptake was modestly higher (10%) in smokers vs. nonsmokers. When subgrouped by sex, regardless of smoking status, [123I]beta-CIT uptake was higher in the striatum (10%), diencephalon (15%), and brainstem (15%) in females vs. males. The sex*smoking interaction was not significant in the striatum, diencephalon, or brainstem, despite the observation of 20% higher brainstem [123I]beta-CIT uptake in male smokers vs. nonsmokers and less than a 5% difference between female smokers and nonsmokers. The results demonstrate higher DA and 5-HT transporter availability in females vs. males and no overall effect of smoking with the exception of a modest elevation in brainstem 5-HT transporters in male smokers. Although these findings are preliminary and need validation with a more selective 5-HT transporter radiotracer, the results suggest that brainstem 5-HT transporters may be regulated by smoking in a sex-specific manner.

Comparison of [(18)F]altanserin and [(18)F]deuteroaltanserin for PET imaging of serotonin(2A) receptors in baboon brain: pharmacological studies.

The regional distribution in brain, distribution volumes, and pharmacological specificity of the PET 5-HT(2A) receptor radiotracer [(18)F]deuteroaltanserin were evaluated and compared to those of its non-deuterated derivative [(18)F]altanserin. Both radiotracers were administered to baboons by bolus plus constant infusion and PET images were acquired up to 8 h. The time-activity curves for both tracers stabilized between 4 and 6 h. The ratio of total and free parent to metabolites was not significantly different between radiotracers; nevertheless, total cortical R(T) (equilibrium ratio of specific to nondisplaceable brain uptake) was significantly higher (34-78%) for [(18)F]deuteroaltanserin than for [(18)F]altanserin. In contrast, the binding potential (Bmax/K(D)) was similar between radiotracers. [(18)F]Deuteroaltanserin cortical activity was displaced by the 5-HT(2A) receptor antagonist SR 46349B but was not altered by changes in endogenous 5-HT induced by fenfluramine. These findings suggest that [(18)F]deuteroaltanserin is essentially equivalent to [(18)F]altanserin for 5-HT(2A) receptor imaging in the baboon.

Reproducibility of in vivo brain measures of 5-HT2A receptors with PET and.

The test/retest reproducibility of brain measures of 5-HT2A receptors with positron emission tomography (PET) and [18F]deuteroaltanserin was examined in a group of eight healthy human subjects. PET measures of 5-HT2A receptors were obtained under an equilibrium paradigm, with a 40-min PET acquisition starting approximately at 300 min (308+/-11 min) after bolus plus constant infusion of the radiotracer. Three brain outcome measures were obtained at equilibrium, V(3) (ratio of specific brain uptake to free parent plasma concentration of radiotracer), V(3)' (ratio of specific brain uptake to total parent plasma concentration) and RT (ratio of specific to non-displaceable brain uptakes). V(3)' and RT had high test/retest reproducibility, as measured by mean intra-subject% change for cortical brain areas of 14.1 and 11.0%, respectively. They also had high reliability, as measured by mean intra-class correlation coefficients (ICC) for cortical brain areas of 0.86 and 0.88, respectively. V(3) had low test/retest reproducibility, due to high variability in the measures of free parent tracer in plasma. This study supports the feasibility of equilibrium imaging of 5-HT2A receptors with PET and [18F]deuteroaltanserin. The equilibrium imaging method with [18F]deuteroaltanserin allows a single acquisition and blood measurement to provide an image whose pixel values equal a receptor volume of distribution. Since the single image pixel values are proportional to receptor densities, the images can be used in pixel-by-pixel statistical methods, such as SPM, to assess the distribution and density of 5-HT2A receptors in neuropsychiatric disorders.

SPECT imaging with the D(4) receptor antagonist L-750,667 in nonhuman primate brain.

The suitability of an (123)I-labeled form of the putative D(4) receptor ligand L750,667 as a radiotracer for single photon emission computed tomography imaging was assessed in nonhuman primates. [(123)I]L750,667, labeled by iododestannylation, was administered to baboons in bolus and bolus plus constant infusion paradigms and imaged for 6 h. Total [(123)I]L750,667 brain uptake peaked (2.3% injected dose) at 15 min postinjection. [(123)I]L750,667 uptake was observed in all brain regions measured including diencephalon, brainstem, basal ganglia, cingulate cortex, and cerebellum, and slightly lower levels were noted in the frontal, parietal, temporoinsular, and occipital cortices. Administration of the D(4) receptor antagonist NGD 94-1 (2 mg/kg) did not displace radioactivity from any of the brain regions examined. Thus, while L750,667 is selective for the D(4) receptor in vitro, because brain [(123)I]L750,667 uptake was not displaced by NGD 94-1 at receptor saturating doses, [(123)I]L750,667 does not appear to be a suitable radiotracer for in vivo imaging of the D(4) receptor.

Prediction of dopamine transporter binding availability by genotype: a preliminary report.

OBJECTIVE: Evidence of a relationship between genotype and binding availability was assessed for the dopamine and serotonin transporter genes. METHOD: The authors assessed dopamine transporter genotype at the SLC6A3 3' variable number of tandem repeats (VNTR) polymorphism and serotonin transporter genotype at the SLC6A4 promotor VNTR polymorphism in 30 healthy subjects who also underwent single photon emission computed tomography with [(123)I]beta-CIT. RESULTS: Subjects homozygous for the 10-repeat allele at the SLC6A3 locus demonstrated significantly lower dopamine transporter binding than carriers of the nine-repeat allele. There was no effect of SLC6A4 genotype upon serotonin transporter binding. CONCLUSIONS: These findings suggest that genetic variation at the SLC6A3 3' VNTR polymorphism may modify dopamine transporter function.

Elevated central serotonin transporter binding availability in acutely abstinent cocaine-dependent patients.

OBJECTIVE: Recent work has underscored the role of serotonergic neurotransmission in chronic neural adaptations to cocaine dependence. The authors tested for evidence of serotonergic dysfunction during acute abstinence from cocaine, a period of high risk for relapse in cocaine dependence. METHOD: Binding availability of dopamine transporters and serotonin transporters was measured in 15 cocaine-dependent subjects during acute abstinence and in 37 healthy comparison subjects by using [(123)I]beta-CIT and single photon emission computed tomography. RESULTS: Significant increases in diencephalic and brainstem serotonin transporter binding (16.7% and 31.6%, respectively) were observed in cocaine-dependent subjects. Brainstem serotonin transporter binding was significantly inversely correlated with age across diagnostic groups. CONCLUSIONS: These findings provide further evidence of serotonergic dysfunction during acute abstinence from chronic cocaine use. Age-related decline in brainstem serotonin transporter binding may underlie the poor response to selective serotonin reuptake inhibitor antidepressants seen in some elderly depressed patients.

Serotonin transporters upregulate with chronic cocaine use.

Cocaine potently inhibits serotonin (5-HT) reuptake in cell bodies and at nerve terminals and 5-HT has been implicated as a modulator of dopaminergic neurotransmission. Chronic use of cocaine may lead to a "serotonin-deficit" form of 5-HT dysregulation. We have examined the status of the 5-HT transporter (SERT) using ligand binding and autoradiographic methods in subgroups of cocaine overdose deaths. Quantitative autoradiography of [125I]RTI-55 was used to map and measure the effect of chronic cocaine use on SERT densities in the striatum, substantia nigra, amygdala, and adjacent paralimbic cortical areas of cocaine overdose (CO) victims with and without preterminal evidence of excited delirium (ED). SERT densities were elevated in the nucleus accumbens and throughout the anterior and posterior sectors of striatum in CO victims compared with age-matched and drug-free control subjects. In contrast, SERT densities were increased significantly in the anterior striatum, but not the posterior sectors in ED victims. Significant elevations in SERT were measured in the orbitofrontal gyrus (Brodmann area 11), the anterior portion of the insular cortex and the cingulate gyrus (Brodmann area 24) in CO and ED victims. Saturation binding site analysis demonstrated an increase in the density of RTI-55 binding sites with no change in the affinity of the radioligand for the SERT. Chronic cocaine exposure upregulated SERT densities in the substantia nigra of the CO, but not ED victims. The lack of SERT upregulation in the substania nigra and posterior striatum suggests the possibility of a distinct phenotype for fatal ED victims that exhibited an acute onset of bizarre and violent behavior prior to death. Adaptive changes in the SERT densities may contribute to depressed mood and drug craving associated with acute cocaine abstinence.

Equilibrium modeling of 5-HT(2A) receptors with [18F]deuteroaltanserin and PET: feasibility of a constant infusion paradigm.

[(18)F]Altanserin has emerged as a promising positron emission tomography (PET) ligand for serotonin-2A (5-HT(2A)) receptors. The deuterium substitution of both of the 2'-hydrogens of altanserin ([(18)F]deuteroaltanserin) yields a metabolically more stable radiotracer with higher ratios of parent tracer to radiometabolites and increased specific brain uptake than [(18)F]altanserin. The slower metabolism of the deuterated analog might preclude the possibility of achieving stable plasma and brain activities with a bolus plus constant infusion within a reasonable time frame for an (18)F-labeled tracer (T(1/2) 110 min). Thus, the purpose of this study was to test the feasibility in human subjects of a constant infusion paradigm for equilibrium modeling of [(18)F]deuteroaltanserin with PET. Seven healthy male subjects were injected with [(18)F]deuteroaltanserin as a bolus plus constant infusion lasting 10 h postinjection. PET acquisitions and venous blood sampling were performed throughout the infusion period. Linear regression analysis revealed that time-activity curves for both specific brain uptake and plasma [(18)F]deuteroaltanserin concentration stabilized after about 5 h. This permitted equilibrium modeling and estimation of V(')(3) (ratio of specific uptake to total plasma parent concentration) and the binding potential V(3) (ratio of specific uptake to free plasma parent concentration). Cortical/cerebellar ratios were increased by 26% relative to those we previously observed with [(18)F]altanserin using similar methodology in a somewhat older subject sample. These results demonstrate feasibility of equilibrium imaging with [(18)F]deuteroaltanserin and suggest that it may be superior to [(18)F]altanserin as a PET radioligand.

Kinetic and equilibrium analyses of [(123)I]epidepride binding to striatal and extrastriatal dopamine D(2) receptors.

Quantitative SPECT measures of dopamine D(2) like receptors with [(123)I]epidepride is complicated by its high affinity and lipophilic metabolites. The purpose of this study was to use both parent (P) and lipophilic metabolites (M) as input functions in a kinetic paradigm and in comparison to the results of equilibrium studies. Kinetic studies on eleven healthy human subjects, ages 32+/- 10 were performed following i.v. injection of approximately 370 MBq of [(123)I]epidepride. Images were acquired for 13.5+/-1.0 hours. Equilibrium studies were done on seven of eleven subjects with a bolus injection of approximately 140 MBq, bolus/infusion ratio of 10 hours, and infusion for 30-32 hours. High (striatum) and low (temporal cortex) density regions were studied. Two (P and M) and one (P) input function models were applied in the kinetic studies. In receptor-rich regions, the distribution volumes in nondisplaceable compartments were fixed to those in cerebellum. In addition, in the two input function model, K(1)(P)/K(1)(M) was fixed to the values in the cerebellum. The one input function model provided V'(3) values (=f(1)*B'(max)/K(D)) which were consistent with those obtained in equilibrium studies in both receptor-rich regions, while the two input function model provided consistent values only in striatum. Poor identifiability of the rate constants of metabolites seemed to be the source of errors in the two input function model. These results suggest that correct V'(3) values can be obtained with the one input function model both in high- and low-density regions.

Characterization of kappa1-opioid receptor binding in human insular cortex.

Mesolimbic dopaminergic neurotransmission is modulated by dynorphin peptides binding to kappa-opioid receptors. The interaction between dynorphin and dopamine systems makes the kappa-opioid receptor a potential drug discovery target for the development of therapeutic agents for schizophrenia and drug abuse. This study reports the specificity and parameters of [3H]U69593 binding in the insular cortex, a representative corticolimbic area of the human brain. The results demonstrate that the radioligand [3H]U69593 labels a single population of receptors in human insular cortex with an affinity in the low nanomolar range. The pharmacological profile for inhibition of [3H]U69593 binding was determined in this brain region using drugs known to bind to mu, kappa and delta opioid receptors. The results show that kappa-opioid selective agonists and antagonists inhibit binding of this ligand in human brain with comparable affinities and rank order as previously described for rat and guinea pig brain and the cloned kappa1-opioid receptor subtype.

D3 dopamine and kappa opioid receptor alterations in human brain of cocaine-overdose victims.

Cocaine is thought to be addictive because chronic use leads to molecular adaptations within the mesolimbic dopamine (DA) circuitry, which affects motivated behavior and emotion. Although the reinforcing effects of cocaine are mediated primarily by blockade of DA uptake, reciprocal signaling between DA and endogenous opioids has important implications for understanding cocaine dependence. We have used in vitro autoradiography and ligand binding to map D3 DA and kappa opioid receptors in the human brains of cocaine-overdose victims. The number of D3 binding sites was increased one-to threefold over the nucleus accumbens and ventromedial sectors of the caudate and putamen from cocaine-overdose victims, as compared to age-matched and drug-free control subjects. D3 receptor/cyclophilin mRNA ratios in the nucleus accumbens were increased sixfold in cocaine-overdose victims over control values, suggesting that cocaine exposure also affects the expression of D3 receptor mRNA. The number of kappa opioid receptors in the nucleus accumbens and other corticolimbic areas from cocaine fatalities was increased twofold as compared to control values. Cocaine-overdose victims exhibiting preterminal excited delirium had a selective upregulation of kappa receptors measured also in the amygdala. Understanding the complex regulatory profiles of DA and opioid synaptic markers that occur with chronic misuse of cocaine may suggest multitarget strategies for treating cocaine dependence.

Immunocytochemical localization of the dopamine transporter in human brain.

The dopamine transporter (DAT) was localized in normal human brain tissue by light microscopic immunocytochemistry by using highly specific monoclonal antibodies. Regional distribution of DAT was found in areas with established dopaminergic circuitry, e.g., mesostriatal, mesolimbic, and mesocortical pathways. Mesencephalic DAT-immunoreactivity was enriched in the dendrites and cell bodies of neurons in the substantia nigra pars compacta and ventral tegmental area. Staining in the striatum and nucleus accumbens was dense and heterogeneous. Mesocortical DAT immunoreactivity in motor, premotor, anterior cingulate, prefrontal, entorhinal/perirhinal, insular, and visual cortices was detected in scattered varicose and a few nonvaricose fibers. Varicose fibers were relatively enriched in the basolateral and central subnuclei of amygdala, with sparser fibers in lateral and basomedial subnuclei. Double-labeling studies combining DAT and tyrosine hydroxylase (TH) immunostaining in the ventral mesencephalon showed two subpopulations of dopaminergic neurons differentiated by the presence or absence of DAT-immunoreactivity in the A9 and A10 cell groups. In other dopaminergic cell groups (All, A13-A15), TH-positive hypothalamic neurons showed no detectable DAT-immunoreactivity. However, fine DAT-immunoreactive axons were scattered throughout the hypothalamus, particularly concentrated along the medial border, with more coarse axons present along the lateral border. These findings demonstrate that most mesotelencephalic dopamine neurons of human brain express high levels of DAT throughout their entire somatodendritic and axonal domains, whereas a smaller subpopulation of mesencephalic dopamine cells and all hypothalamic dopamine cell groups examined express little or no DAT. These data indicate that different subpopulations of dopaminergic neurons use different mechanisms to regulate their extracellular dopamine levels.

Imaging of the serotonergic system: interactions of neuroanatomical and functional abnormalities of depression.

For nearly three decades, evidence supporting a role for aberrant serotonergic function in the pathogenesis of depression has accumulated; however, only recently have methodologies and radiotracers suitable for in vivo clinical assessment of depression become available. To date, only a few neurochemical imaging studies have been performed in actively depressed subjects. A preliminary study using single photon emission computed tomography (SPECT) has demonstrated decreased levels of serotonin (5-HT) transporters in the midbrain regions of subjects with major depression. Analysis of the 5-HT2 receptor using positron emission tomography (PET) has suggested that this receptor may not be altered significantly in the depressed brain but may increase in response to antidepressant treatment. These findings are supported by studies in secondary "poststroke" depression that have shown that elevations in 5-HT2 receptor density correlated with the alleviation of symptoms of depressed mood. With the rapid development of novel PET and SPECT radiotracers, future studies of the serotonergic system that evaluate presynaptic (5-HT transporter) and postsynaptic (5-HT1A and 5-HT2A receptors) markers and the interaction of synaptic levels of 5-HT with these sites will make profound contributions to the understanding of the role of the serotonergic synapse in the pathophysiology of depression.

Characterization and localization of galanin receptors in human entorhinal cortex.

The neuropeptide galanin (GAL) has a widespread distribution throughout the human cortex. The entorhinal cortex (ENT) plays a crucial role in the transfer of cortico-cortical information related to memory and displays severe degeneration in Alzheimer's disease (AD). However, very little is known about the pharmacology of the GAL receptor (GALR) in normal human ENT. Therefore, we pharmacologically visualized their distribution and characterized GALRs using in vitro receptor autoradiography and radioligand binding assays. Autoradiograms revealed intense GALR labeling, mainly in the substantia innominata, hypothalamus, the bed nucleus of the stria terminalis and within layers 2 and 4 of the ENT. Kinetic experiments showed that saturation of GALR sites by [125I]GAL (human) (hGAL) occurred within 2 h and that this binding readily reversed in the presence of a GTP analog, but not in the presence of excess unlabeled hGAL. Analysis of [125I]hGAL binding data from saturation experiments gave KD values of 98.6+/-21.6 pM, Bmax values of 52.9+/-32.4 fmol/mg protein and identified a high and low affinity state of the GALR. The presence of 5'-guanylylimidodiphosphate (GppNHp) or NaCl reduced the agonist labeling of hGALR in ENT membranes.

Kappa2 opioid receptors in limbic areas of the human brain are upregulated by cocaine in fatal overdose victims.

Cocaine is thought to be addictive because chronic use leads to molecular adaptations within the mesolimbic dopamine (DA) circuitry that affect motivated behavior and emotion. Although the reinforcing effects of cocaine are mediated primarily by blocking DA reuptake into the presynaptic nerve terminal, reciprocal signaling between DA and endogenous opioids has important implications for cocaine dependence. The present study used the opioid antagonist 6 beta-[125iodo]-3,14-dihydroxy-17-cyclopropylmethyl-4,5 alpha-epoxymorphinan ([125I]IOXY) after pretreatment with the site-directed acylating agents 2-(p-ethoxybenzyl)-1-diethylaminoethyl-5-isothiocyanatobenzimid iazole -HCl (mu-selective) and N-phenyl-N-[1-(2-(4-isothiocyanato)-phenethyl)-4-piperidinyl]-p ropana mide-HCl (delta-selective) to examine the effect of cocaine exposure on the distribution and density of kappa2 receptors in autopsy studies of human cocaine fatalities. The selective labeling of the kappa2 receptor subtype was demonstrated by competition binding studies, which gave a pharmacological signature (IOXY >/= (+)-bremazocine >> U50,488 >/= U69,593) distinct from either the kappa1 or kappa3 receptor subtypes. Visualization of [125I]IOXY labeling revealed that kappa2 receptors localize to mesocortical and subcortical limbic areas, including the cingulate, entorhinal, insular, and orbitofrontal cortices and the nucleus accumbens and amygdala. The number of kappa2 receptors in the nucleus accumbens and other limbic brain regions from cocaine fatalities was increased twofold as compared with age-matched and drug-free control subjects. Cocaine overdose victims, who experienced paranoia and marked agitation before death, also had elevated densities of kappa2 receptors in the amygdala. These findings demonstrate for the first time that kappa2 receptor numbers are upregulated by cocaine exposure. The molecular adaptation of kappa2 receptor numbers may play a role in the motivational incentive associated with episodes of binge cocaine use and in the dysphoria that follows abrupt cocaine withdrawal.

Vesicular acetylcholine transporter density and Alzheimer's disease.

We have evaluated the vesamicol analogue meta-[125I]iodobenzyltrozamicol {(+)-[125I]MIBT} as a probe to assess cholinergic terminal integrity in the human temporal cortex. Saturation binding analysis, using 5-aminobenzovesamicol (ABV) to define nonspecific binding, revealed a high-affinity binding site with a Kd value of 4.3 +/- 1.2 nM in the temporal cortex of the young control subjects. Similar affinity values were observed for (+)-[125I]MIBT binding in aged control subjects (Kd = 3.4 +/- 0.5 nM) and AD patients (Kd = 3.0 +/- 0.8 nM). In contrast, Bmax values for young subjects, aged controls and AD patients were 31.2 +/- 6.3, 17.0 +/- 2.0 and 9.4 +/- 1.6 pmol/g, respectively, clearly reflecting significant reductions in (+)-[125I]MIBT binding site density with aging and age-related neuropathology. Moreover, the decrease in (+)-[125I]MIBT binding was correlated with choline acetyltransferase activities (r = 0.72) in the AD temporal cortex. These results suggest that when selective ligands are used, the vesicular acetylcholine transporter can be a useful marker protein for assessing the loss of cholinergic projections in AD and related disorders.

Immunochemical analysis of dopamine transporter protein in Parkinson's disease.

The plasma membrane dopamine transporter (DAT) is considered to be a reliable marker of presynaptic dopaminergic terminal loss. Previous in vivo imaging and postmortem binding studies have detected a loss in striatal DAT binding in Parkinson's diseased (PD) brain; however, these techniques have poor spatial resolution and may suffer from nonspecific binding of some ligands. In this study, we use novel highly specific monoclonal antibodies to distinct epitopes of human DAT to quantify and localize the protein. Western blot analysis revealed marked reductions in DAT immunoreactivity in putamen, caudate, and nucleus accumbens of PD brain compared with control cases, and the reductions were significantly correlated to disease duration. Immunohistochemistry revealed DAT-immunoreactive fibers and puncta that were dense throughout the striatum of control brains but that were drastically reduced in putamen of PD brains. Caudate from PD brains showed a significant degree of sparing along the border of the ventricle, and the nucleus accumbens was relatively preserved. An unexpected finding was that discrete islands of DAT immunoreactivity were preserved within the matrix of PD putamen. Thus, immunological analysis of DAT protein provides novel and sensitive means for localizing and quantifying DAT protein in PD and other neurological disorders involving dopaminergic systems.

Radioligand binding and immunoautoradiographic evidence for a lack of toxicity to dopaminergic nerve terminals in human cocaine overdose victims.

Radioligand binding to and immunolabeling of transport sites associated with monoamine-containing synaptic vesicles affords a novel approach for mapping the integrity of dopaminergic (DAergic) nerve terminals. The present study used [125I]iodovinyltetrabenazine ([125I]TBZ) and a fusion protein antibody directed at the large intraluminal loop of the neuronal vesicular monoamine transporter (hVMAT2-loop) as probes to assess the effects of chronic cocaine use on the integrity of DAergic nerve terminals in the striatum of cocaine fatalities. Visualization of [125I]TBZ binding in human brain revealed a distinct pattern of labeling throughout the rostral-caudal extent of the striatum. Saturation binding of [125I]TBZ in striatal membranes demonstrated a single high affinity site (Kd = 2.3 +/- 0.9 nM and Bmax = 55.5 +/- 8.1 pmol/g tissue) with a pharmacological profile (tetrabenazine > or = iodovinyltetrabenazine > ketanserin > or = reserpine > haloperidol > GBR 12909) consistent with the specific labeling of hVMAT2. Quantitative in vitro autoradiography demonstrated no significant alteration in the density of [125I]TBZ binding sites in the anterior and posterior sectors of the striatum in cocaine fatalities with and without preterminal excited delirium as compared to drug-free and age-matched control subjects. Similarly, the levels of hVMAT2-loop immunoreactivity were not significantly different across control and cocaine fatality groups. The results demonstrate the lack of an alteration in [125I]TBZ binding sites and hVMAT2 protein in the striatum from a young cohort of cocaine fatalities. Since striatal VMAT2 is primarily associated with DAergic nerve terminals, these results suggest that chronic cocaine use failed to affect the integrity of striatal DAergic nerve terminals.

Pharmacological characterization of the vesamicol analogue (+)-[(125)I]MIBT in primate brain.

The vesamicol analogue, meta-[(125)I]iodobenzyltrozamicol [(+)-[(125)I]MIBT] was evaluated as a probe for the in vitro labeling of the vesicular acetylcholine transporter in primate brain. In the striatum, (+)-[(125)I]MIBT bound a single high-affinity site with a Kd value of 4.4 +/- 0.7 nM. Competition for (+)-[(125)I]MIBT binding to the striatum by a group of vesamicol analogues displayed a pharmacological profile similar to the rank order of potency previously observed for the vesicular acetylcholine transporter on Torpedo synaptic vesicles. High-affinity binding of (+)-[(125)I]MIBT in the occipital cortex was characterized by a Kd value of 4.6 +/- 1.1 nM. However, the rank order of potency for inhibition of (+)-[(125)I]MIBT binding to the occipital cortex by the same test compounds differed from that observed in the striatum. The results suggest that (+)-[(125)I]MIBT is a reliable probe of the vesicular acetylcholine transporter in primate striatum, but its binding in primate occipital cortex is more complex.

Adaptive increase in D3 dopamine receptors in the brain reward circuits of human cocaine fatalities.

The mesolimbic dopaminergic system plays a primary role in mediating the euphoric and rewarding effects of most abused drugs. Chronic cocaine use is associated with an increase in dopamine neurotransmission resulting from the blockade of dopamine uptake and is mediated by the activation of dopamine receptors. Recent studies have suggested that the D3 receptor subtype plays a pivotal role in the reinforcing effects of cocaine. The D3 receptor-preferring agonist 7-hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT) is a reinforcer in rhesus monkeys trained to self-administer cocaine, but not in cocainenaive monkeys. In vitro autoradiographic localization of [3H]-(+)-7-OH-DPAT binding in the human brain demonstrated that D3 receptors were prevalent and highly localized over the ventromedial sectors of the striatum. Pharmacological characterization of [3H]-(+)-7-OH-DPAT binding to the human nucleus accumbens demonstrated a rank order of potency similar to that observed for binding to the cloned D3 receptor expressed in transfected cell lines. Region-of-interest analysis of [3H]-(+)-7-OH-DPAT binding to the D3 receptor demonstrated a one- to threefold elevation in the number of binding sites over particular sectors of the striatum and substantia nigra in cocaine overdose victims as compared with age-matched and drug-free control subjects. The elevated number of [3H]-(+)-7-OH-DPAT binding sites demonstrates that adaptive changes in the D3 receptor in the reward circuitry of the brain are associated with chronic cocaine abuse. These results suggest that the D3 receptor may be a useful target for drug development of anticocaine medications.