Sudha Garg

Kinetics of brain nicotine accumulation in dependent and nondependent smokers assessed with PET and cigarettes containing 11C-nicotine

Tobacco smoking is a chronic, relapsing disorder that constitutes one of the primary preventable causes of death in developed countries. Two of the popular hypotheses to explain the development and maintenance of strong nicotine dependence in cigarette smokers posit (i) a rapid brain nicotine accumulation during cigarette smoking and/or (ii) puff-associated spikes in brain nicotine concentration. To address these hypotheses, we investigated the dynamics of nicotine accumulation in the smokers brain during actual cigarette smoking using PET with 3-s temporal resolution and 11C-nicotine loaded into cigarettes. The results of the study, performed in 13 dependent smokers (DS) and 10 nondependent smokers (NDS), suggest that puff-associated spikes in the brain nicotine concentration do not occur during habitual cigarette smoking. Despite the presence of a puff-associated oscillation in the rate of nicotine accumulation, brain nicotine concentration gradually increases during cigarette smoking. The results further suggest that DS have a slower process of brain nicotine accumulation than NDS because they have slower nicotine washout from the lungs and that DS have a tendency to compensate for their slower rate of brain nicotine accumulation compared with NDS by inhaling a larger volume of smoke. For these reasons, smokers’ dependence on cigarette smoking, or the resistance of NDS to becoming dependent, cannot be explained solely by a faster brain nicotine accumulation.

Effect of Menstrual Cycle Phase on Dopamine D2 Receptor Availability in Female Cynomolgus Monkeys

Sex differences have been reported in a variety of affective and neurodegenerative disorders that involve dysfunctional dopamine (DA) neurotransmission. In addition, there is evidence for differences in sensitivity to the abuse-related effects of psychostimulants across the menstrual cycle which may result from effects of ovarian hormones on DA function. The goal of the present study was to extend previous work examining menstrual cycle-related changes in DA D2 receptor availability in humans to drug-naive female cynomolgus monkeys (n=7) using the selective D2-like receptor ligand [18F]fluoroclebopride (FCP) and a high-resolution microPET P4 scanner. Menstrual cycle phase was characterized by daily vaginal swabs and measurements of serum progesterone levels. PET studies were conducted once during the luteal phase and once during the follicular phase. Regions of interest in the caudate nucleus, putamen, and cerebellum were defined on coregistered MRIs. Distribution volumes were calculated for FCP in each structure and the distribution volume ratio (DVR) for both brain regions relative to the cerebellum was used as a measure of D2 receptor availability. FCP DVRs were significantly higher in the luteal phase compared to the follicular phase in both the caudate nucleus (11.7% difference, p=0.02) and putamen (11.6% difference, p=0.03). These findings extend earlier work in humans and suggest that changes in DA receptor availability may be involved in the variation in symptoms of various neuropsychiatric disorders across the menstrual cycle, including differences in sensitivity to the abuse-related effects of stimulants.

Effects of Cocaine and MDMA Self-Administration on Serotonin Transporter Availability in Monkeys

Although serotonin (5-HT) can interact with dopamine (DA) systems to modulate the subjective and reinforcing effects of psychostimulants such as cocaine and 3,4-methyldioxymethamphetamine (MDMA, ecstasy), the long-term effects of exposure to psychostimulants on brain 5-HT systems are not well characterized. The present study assessed 5-HT transporter (SERT) availability using positron emission tomography (PET) in rhesus monkeys with the SERT-specific radioligand [11C]3-amino-4-(2-dimethylaminomethyl-phenylsulfanyl)-benzonitrile (DASB). SERT availability was assessed in regions of interest including the caudate nucleus, putamen, anterior cingulate cortex, and cerebellum. [11C]DASB distribution volume ratios (DVRs) were calculated using the cerebellum as the reference region. DVRs were calculated in control monkeys and in cocaine or MDMA self-administering monkeys approximately 24 h after the last self-administration (SA) session. SERT availability did not differ between monkeys with a history of MDMA SA and control monkeys in any region examined. In contrast, monkeys with a history of cocaine SA showed significantly higher levels of SERT availability in the caudate nucleus and putamen compared to control subjects. These results suggest that chronic SA of cocaine, but not MDMA, leads to alterations in serotonergic function in brain areas relevant to drug abuse. The higher level of SERT availability in cocaine-experienced monkeys may lead to a reduced inhibitory tone of 5-HT on the DA system, which may explain, in part, differences in the abuse liability between cocaine and MDMA.

Social Dominance in Female Monkeys: Dopamine Receptor Function and Cocaine Reinforcement

BACKGROUND Brain imaging and behavioral studies suggest an inverse relationship between dopamine (DA) D2/D3 receptors and vulnerability to cocaine abuse, although most research has used males. For example, male monkeys that become dominant in a social group have significant elevations in D2/D3 receptor availability and are less vulnerable to cocaine reinforcement. METHODS DA D2/D3 receptor availability was assessed in female cynomolgus monkeys (n = 16) with positron emission tomography (PET) while they were individually housed, 3 months after stable social hierarchies had formed, and again when individually housed. In addition, PET was used to examine changes in dopamine transporter (DAT) availability after social hierarchy formation. After imaging studies were complete, monkeys received implantation with indwelling intravenous catheters and self-administered cocaine (.001-.1 mg/kg/injection) under a fixed-ratio 30 schedule of reinforcement. Acquisition of cocaine reinforcement occurred when response rates were significantly higher than when saline was self-administered. RESULTS Neither DAT nor D2/D3 receptor availability in the caudate nucleus and putamen was predictive of social rank, but both significantly changed after formation of social hierarchies. DA D2/D3 receptor availability significantly increased in females that became dominant, whereas DAT availability decreased in subordinate females. Dominant female monkeys acquired cocaine reinforcement at significantly lower doses than subordinate monkeys. CONCLUSIONS The relationship between D2/D3 receptor availability and vulnerability to cocaine reinforcement seems, on the basis of these findings, opposite in females and males. These data indicate that the social environment profoundly affects the DA system but does so in ways that have different functional consequences for females than for males.

Behavioral and neurobiological characteristics influencing social hierarchy formation in female cynomolgus monkeys.

Socially housed monkeys have been used as a model to study human diseases. The present study examined behavioral, physiological and neurochemical measures as predictors of social rank in 16 experimentally naïve, individually housed female cynomolgus monkeys (Macaca fascicularis). The two behavioral measures examined were novel object reactivity (NOR), as determined by latency to touch an opaque acrylic box placed in the home cage, and locomotor activity assessed in a novel open-field apparatus. Serum cortisol concentrations were evaluated three times per week for four consecutive weeks, and stress reactivity was assessed on one occasion by evaluating the cortisol response to adrenocorticotropic hormone (ACTH) following dexamethasone suppression. Measures of serotonin (5-HT) function included whole blood 5-HT (WBS) concentrations, cerebrospinal fluid (CSF) concentrations of the 5-HT metabolite 5-hydroxyindoleacetic acid (5-HIAA) and brain 5-HT transporter (SERT) availability obtained using positron emission tomography (PET). After baseline measures were obtained, monkeys were assigned to four social groups of four monkeys per group. The two measures that correlated with eventual social rank were CSF 5-HIAA concentrations, which were significantly higher in the animals who eventually became subordinate, and latency to touch the novel object, which was significantly lower in eventual subordinate monkeys. Measures of 5-HT function did not change as a consequence of social rank. These data suggest that levels of central 5-HIAA and measures of novel object reactivity may be trait markers that influence eventual social rank in female macaques.

Design, Synthesis, and Preliminary in Vitro and in Vivo Evaluation of N-(2-diethylaminoethyl)-4-[18F]fluorobenzamide ([18F]-DAFBA): A Novel Potential PET Probe to Image Melanoma Tumors

In order to develop a PET radiopharmaceutical to image malignant melanoma, we synthesized N-(2-diethylaminoethyl)-4-[(18)F]fluorobenzamide ([(18)F]-DAFBA). In vitro studies show a high uptake of [(18)F]-DAFBA by the B16F1 melanoma cells. No significant binding was seen for DAFBA to the sigma-1 and sigma-2 receptors in vitro. The in vivo biodistribution studies performed in normal ICR mice showed a low uptake in the normal tissues followed by further elimination of radioactivity from these tissues with time. The biodistribution studies performed in C57 mice bearing the melanoma tumor xenograft showed a rapid uptake of radioactivity in the tumor that reached a plateau within 30 min postinjection. The F-18 uptake in the tumor was 7.00 +/- 2.76, 6.57 +/- 1.66, and 5.80 +/- 0.98%ID/g at 60, 120, and 180 min, respectively. A steady uptake of radioactivity in the tumor and low uptake in normal tissues resulted in high tumor to normal tissue ratios. For example, at 180 min postinjection, the tumor to tissue ratios were 14.90 +/- 6.47, 21.90 +/- 4.68, 32.91 +/- 6.11, 39.73 +/- 11.78, and 6.33 +/- 1.9, for the spleen, lungs, muscle, blood, and liver, respectively. The radioactivity rapidly cleared from the blood pool, and it decreased from 0.68 +/- 0.21%ID/g at 60 min to 0.13 +/- 0.03%ID/g at 180 min. The F-18 uptake in the bones at 60, 120, and 180 min was 0.91 +/- 0.27, 0.57 +/- 0.32, and 0.17 +/- 0.05%ID/g, respectively. This low uptake in the bones reflects its in vivo resistance toward defluorination. A low residual activity in normal tissues and a high tumor uptake signifies the superior imaging potential of this compound. Because of these positive traits, [(18)F]-DAFBA could help delineate the tumor and its metastases when used for imaging applications. Further in vivo studies are underway to assess the potential of [(18)F]-DAFBA as a promising PET imaging probe.

Synthesis and preliminary evaluation of para- and meta-[18F]fluorobenzylguanidine.

meta-[18F]Fluorobenzylguanidine ([18F]MFBG) and para-[18F]fluorobenzylguanidine ([18F]PFBG) were synthesized in three steps beginning with a fluoro for nitro exchange reaction on 3- and 4-nitrobenzonitrile, respectively. Overall radiochemical yields were 10-15% for [18F]MFBG and 50-55% for [18F]PFBG in a total synthesis time of 60 min. However, impurities interfered with the binding of the product to target cells. A new route was adopted for the synthesis of [18F]PFBG using 4-nitrilophenyl trimethylammonium trifluoromethanesulfonate (Q.S.) as the starting material. In addition to shortening the overall synthesis time by 10 min, this precursor also eliminated problems associated with the presence of small amounts of starting material in the preparation. In vitro binding of [18F]PFBG prepared by the Q.S. method to SK-N-SH, human neuroblastoma cells was 26.5 +/- 1.1%, compared to 16.9 +/- 1.6% when the nitro precursor was used. Selective uptake of both 18F-labeled isomers in the heart and adrenal was seen in mice. At 4 h, adrenal and heart uptake of [18F]PFBG prepared using Q.S. was 20.3 +/- 4.8 and 5.9 +/- 0.8% ID/g respectively, compared to 23.8 +/- 5.0 and 10.5 +/- 1.7% ID/g for [18F]MFBG. Based on the 5-fold higher radiochemical yields obtained with [18F]PFBG, this isomer would appear to be the more practical choice; however, in vitro and in vivo results suggest that [18F]MFBG exhibits greater similarities to MIBG.

Radioiodination of a monoclonal antibody using N-succinimidyl 5-iodo-3-pyridinecarboxylate

The potential utility of N-succinimidyl 5-iodo-3-pyridinecarboxylate (SIPC) for the radioiodination of monoclonal antibodies was investigated. Paired-label studies were performed using the anti-tenascin antibody 81C6 in athymic mice bearing subcutaneous D-54 MG human glioma xenografts. Radiolabeling was also done using N-succinimidyl 3-iodobenzoate (SIB). Radioiodination of SIPC and SIB both proceeded in 60-80% yield, but protein coupling efficiencies with SIB were higher (76 +/- 16 vs 60 +/- 7%). Immunoreactivity and affinity of both preparations were similar. Using SIPC, thyroid uptake was quite low, decreasing from 0.3% at day 1 to 0.05% at day 8. Tumor uptake reached 46 +/- 11% injected dose/g at day 1 but declined gradually thereafter. This apparent decline reflected the rapid growth of these xenografts since tumor accumulation expressed as percentage of injected dose remained nearly constant up to day 9. These results suggest that SIPC, like SIB, offers significant advantages for labeling antibodies when compared with conventional protein iodination methods.

Sex-specific effects of cigarette mentholation on brain nicotine accumulation and smoking behavior.

Menthol cigarettes are likely associated with greater risks of smoking dependence than non-menthol cigarettes. We sought to test the hypothesis that menthol increases the rate of brain nicotine accumulation (BNA) during smoking and thereby enhances its addictive effects. In a counter-balanced cross-over design, 10 menthol and 9 non-menthol smokers (10 females and 9 males; mean age 44.3) underwent two study phases. In each phase, the participant smoked exclusively either menthol or non-menthol research cigarettes for approximately 1 week prior to a positron emission tomography (PET) scan session, during which the subject’s head was scanned following inhalation of a single puff of smoke from a cigarette containing 11C-nicotine. No differences in initial slope, Cmax, area under curve (AUC), and T1/2 of BNA were found between menthol and non-menthol cigarettes across all subjects; however, menthol relative to non-menthol cigarettes were associated with steeper initial slopes in men (p=0.008). Unexpectedly, women had faster BNA as indicated by greater values of the initial slope, Cmax, AUC, and shorter T1/2 than men (all ps<0.04). The rates of BNA were significantly correlated with ratings of smoking motivations of getting a ‘rush’, getting relaxing effects and marginally with alleviation of craving. These results do not provide strong support for the putative role of menthol in enhancing BNA, although further studies should explore the apparent effect of menthol on BNA in men. Fast BNA during smoking and preference of sensory properties of menthol cigarettes may independently or jointly contribute to smoking dependence among women.

Para-[18F]fluorobenzylguanidine kinetics in a canine coronary artery occlusion model

BackgroundThe kinetics of para-[18F]fluorobenzylguanidine ([18F]PFBG) were investigated in a canine coronary artery occlusion model.Methods and ResultsFive dogs were imaged by positron emission tomography (PET) before and after complete surgical ligation of the left anterior descending coronary artery. PET studies included a 10-minute dynamic [13N]NH3 perfusion scan, followed 1 hour later by 3-hour dynamic [18F]PFBG scanning. [18F]PFBG and [13N]NH3 images demonstrated homogeneous myocardial uptake/perfusion before infarction. One hundred eighty minutes after [18F]PFBG administration, myocardial accumulation was decreased by 60% (day, 2, 0.0065% ±0.0015% injected dose/ml) and 58% (day 16, 0.0069%±0.003% injected dose/ml) compared with a similar myocardial region of interest from the preinfarction (0.016%±0.005% injected dose/ml) study. Myocardial accumulation of [13N]NH3 at 9 minutes showed a 52% (day 2) and 7% (day 16) decrease compared with the preinfarction study. The accumulation of [18F]PFBG in the infarction was decreased significantly at 120 and 180 minutes on all postinfarction studies (p=0.01). In three dogs a significant decrease in the myocardial norepinephrine concentration was documented in the area of infarction (237±94 ng/gm) versus the noninfarcted (1018±48 ng/gm) myocardium (p=0.001).ConclusionsA decreased accumulation of [18F]PFBG was observed in the area of myocardial infarct in this canine model. The magnitude of the decrease in [18F]PFBG was larger than that seen with [13N]NH3 on day 16 after infarction suggesting reperfusion and persistent sympathetic neuronal dysfunction.