Director's Report to the National Advisory Council on Drug Abuse
February, 1996
Research Findings
Intramural Research
Treatment Branch, Clinical Trials Section
Drs. Kenzie L. Preston, Kenneth Silverman, Annie Umbricht-Schneiter, Anna DeJesus, Ivan D. Montoya, and Charles R. Schuster recently completed a study in which the efficacy of contingency management for improving compliance with naltrexone treatment was evaluated. The preliminary analyses suggest that the availability of the vouchers maintained subjects in treatment, and that vouchers given contingent on naltrexone ingestion selectively increased compliance with naltrexone administration. Thus, contingency management appears to be a useful treatment tool for increasing compliance with naltrexone maintenance in patients who are otherwise poorly motivated. This work, combined with other Clinical Trials research to develop regimens to initiate naltrexone administration during short opiate detoxification, should provide a basis for effective alternatives to opiate agonist maintenance.
Neuroscience Branch, Neuroimaging and Drug Action Section
An overview of the methods for imaging brain structure and metabolic activity using PET and SPECT, and MRI to determine the anatomical substrates of the effects of drug of abuse is presented. The effects of a variety of drugs of abuse, including alcohol, stimulants, benzodiazepines, opioids, and marijuana on these measures are reviewed. JM Stapleton and ED London. Imaging Techniques. In: Encyclopedia of Drugs & Alcohol, Vol. 2, JH Jaffe, ed., Macmillan, New York, NY., pp. 573-576, 1995.
The effectiveness of different nitric oxide synthase inhibitors (L-Nitroarginine and L-Nitroarginine methyl ester, and N-momomethyl-L-arginine) in attenuating opiate withdrawal signs was tested in rats, using a variety of regimens. L-Nitroarginine was more potent that
L-Nitroarginine methylester, and N-momomethyl-L-arginine attenuated some behaviors not affected by the other agents. The potential utility of these agents in the clinical treatment of opiate withdrawal is discussed. London ED, AS Kimes, DB Vaupel. Inhibitors of Nitric Oxide Synthase and the Opioid Withdrawal Syndrome. NIDA Res. Monogr. 147: pp. 170-181, 1995.
Experiments performed on dissociated rat cortical cell cultures examined how ascorbic acid alters the neurotoxic effects of two agents, ascorbic acid and nitric oxide (generated from the breakdown of sodium nitroprusside). Ascorbic acid enhanced toxicity of nitric oxide, but it reduced that of NMDA. The results indicate that ascorbic acid produces neuroprotection by an action at the NMDA receptor, probably by antagonizing Ca 2+ influx starting the cascade of biochemical events that lead to the production of NO. JA Bell, C Beglan, and ED London. Interaction of Ascorbic Acid with the Neurotoxic Effects of NMDA and Sodium Nitroprusside. Life Sci., 58: pp. 367 371, 1995.
The neuroanatomical distribution of the dopamine transporter was assessed by immunohistochemical staining. With the use of newly developed polyclonal antisera raised against conjugated peptides corresponding to specific sequences in the dopamine transporter protein, staining was shown to be specific to dopamine neurons and terminals, with virtually no staining in neurons containing transporters for other amines (norepinephrine and serotonin). Dense staining was observed in the basal ganglia, nigro-striatal bundle and lateral habenula. Dopamine cell bodies in the substantia nigra and ventral tegmental area exhibited moderate staining, and distinct fiber staining with a laminar distribution was observed in the cingulate cortex. The functional significance of variations in the staining for dopamine transporter-like immunoreactivity is discussed. Freed C, R Revay, RA Vaughn, E Kriek, S Grant, GR Uhl, and MJ Kuhar. Dopamine Transporter Immunoreactivity in Rat Brain. J. Comp. Neurology, 359: pp. 340-349, 1995.
The distribution of dopamine immunohistochemical staining was examined in brain regions where the presence of dopamine transporter has been controversial with moderate but sparse staining of fibers in the amygdala and olfactory bulb. In contrast, staining of dopaminergic cell bodies in the median eminence was barely detectable despite light staining of fibers in this area. The results provide further support for the hypothesis that different cell groups differ in their expression of dopamine transporter. Revay, R, RA Vaughn, S Grant, and MJ Kuhar, Dopamine Transporter Immunohistochemistry in Median Eminence, Amygdala, and Other Areas of the Rat Brain. Synapse, in press.
The hypothesis that sigma receptors mediate ischemic brain injury was tested using 4-phyenyl-1 (4-phenylbutyl) piperidine (PPBP), a potent sigma receptor ligand. When the drug was administered during induction of transient focal ischemia, it reduced injury volume dramatically and increased somatosensory evoked potentials, a measure of functional recovery. The results indicate that sigma receptors play an important role in ischemic brain injury, and that PPBP can afford protection when administered at the end of an ischemic episode and during reperfusion. H Takahashi, JR Kirsch, K Hashimoto, ED London, R Koehler, and RJ Traystman. PPBP [4 phenyl-l-(4-phenylbutyl) piperidine], a Potent Sigma--Receptor Ligand, Decreases Brain Injury after Transient Focal Ischemia in Cats. Stroke, 26: pp. 1676-1682, 1995.
A stoichiometric analysis of binding to various domains of the N-methyl-D-aspartate (NMDA) receptor was conducted using membranes from various brain regions of rats. The ratio of the density of [3H]CGP39653 binding to [3H]dizocilpine binding was>1 in frontal cortex and hippocampus, 1 in striatum and spinal cord and<1 in cerebellum. When [3H]dichlorokynurenic acid binding was compared to [3H]dizocilpine binding, the ratios were>1 in frontal cortex, hippocampus and striatum, 3 - 4 in cerebellum, and 2 in spinal cord. These observations suggest that a single channel complex may have more than one binding site for NMDA and/or glycine and that the stoichiometry between the binding domains of the NMDA receptor varies regionally. T Matsunaga, AG Mukhin, and ED London. Regionally Distinct Stoichiometry for N-methyl-D aspartate Receptor Domains in Brain. Neuroreport, in press.
Neuroscience Branch, Molecular Psychiatry Section
Dr. Jean Cadet has shown previously that the antioxidant enzyme CuZnsuperoxide dismutase protects against the toxic effects of the drug. In a recent study he has shown that cells that express the proto-oncogen, bcl-2, are protected against the toxic effects of the drug in vitro. Moreover, using flow cytometry, immunofluorescent staining, and DNA electrophoresis, he has shown that METH can cause DNA strand breaks, chromatin condensation, nuclear fragmentation, and DNA laddering. All these changes were prevented by expression of bcl-2. Moreover, scientists within the section have also shown that METH treatment can cause accumulation of cells in the G2 phase of the cell cycle. This is consistent with METH having caused DNA damage since drugs that cause DNA damage are known to cause cell cycle arrest so that DNA damage is not propagated to daughter cells during the process of mitosis. These data provide, for the first time, evidence that METH can cause significant molecular changes that are consistent with the process of apoptosis. These data also open a new line of investigation that will focus on possible direct genetic effects of these drugs.
Preclinical Pharmacology Branch, Psychobiology Section
Studies of benztropine analogs continue to provide interesting leads for a better understanding of how actions at the dopamine transporter may or may not be translated into physiological effects that result in drug abuse. These compounds will provide interesting leads in the development of treatments for cocaine abuse. Recently commercial interest has been expressed for several of the compounds covered by a U.S. Provisional Patent Application (015,280-237,000) which was filed in June, 1995.
The D3 dopamine receptor system has been proposed as critically involved in cocaine abuse. Unfortunately, there have not been adequate in vivo assays to characterize the effects of D3 agonists, and distinguish them from other drugs acting on the dopamine system. We have developed a behavioral procedure that distinguishes selective D3 agonists from other drugs acting on the dopamine system. This procedure will allow for full characterization of the in vivo pharmacology of this class of novel dopamine receptor agonists.
Cocaine binding curves have two components (high and low affinity) and the inhibition of dopamine uptake can exhibit two components. One of these components, comprising approximately 25% of the total dopamine uptake, exhibits a high sensitivity to cocaine and is inhibited by low concentrations of the drug. The other component is much less sensitive to inhibition. Researchers in the section have recently shown that meperidine, an atypical opioid agonist that shares some structural features with the phenyltropane (WIN) analogs of cocaine, selectively inhibits only the high affinity component of dopamine uptake. This effect is mediated by the dopamine transporter and is not produced by opioid mechanisms. Further, meperidine, in the presence of naltrexone to block its prepotent opioid actions, produces subjective effects like those of cocaine in primates trained to discriminate cocaine from saline. These data suggest that the actions of meperidine that are atypical of opioids are due at least in part to activity at the dopamine transporter. In addition, meperidine appears to interact predominantly with the high affinity component of the dopamine transporter, and this high-affinity component may be the site of importance for the production of cocaine's behavioral effects.
In an attempt to discover a selective dopamine D1 antagonist that would not penetrate the blood brain barrier, a novel class of 3-alkylamino- and 3-cinnamylamino-substituted benzazepine analogs were prepared. Several of these compounds bound with moderately high affinity and selectivity to the dopamine D1 receptor and were D1 antagonists as evidenced by their ability to inhibit dopamine-stimulated adenylyl cyclase. Structure-activity relationships derived from these series of compounds coupled with molecular modeling studies suggested that an amine-accepting binding domain that exists approximately 8 Å away from the pharmacophoric benzazepine nitrogen appears to be important in the binding of the most active compounds. This binding domain had not been previously described and may provide a target for novel, therapeutically useful dopamine D1 antagonists.
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