The 2003 Frontiers in Addiction Research mini-convention brought together participants from diverse scientific disciplines to share advances and discuss future directions in the neuroscience of drug abuse and related areas. Many of the discoveries hold great promise for helping NIDA achieve its goal: to significantly reduce the health and social consequences of drug abuse and addiction throughout the United States. The application of NIDA-supported neuroscience research will enable even greater advances vital to reducing drug abuse, addiction, and their related consequences.
Endocannabinoids in the Brain: From Micro to Macro
During this section of the program, presenters discussed the central mechanisms that mediate activities of the endogenous and exogenous cannabinoids, as well as the functional impact of these activities.
- Cross-Talk Between Cannabinoid and Opiate Systems
Steven R. Goldberg, Ph.D., NIDA Intramural Research Program
Dr. Steven Goldberg described a series of studies demonstrating that both THC, the main psychoactive ingredient in marijuana, and anandamide, an endogenous cannabinoid, can serve as effective reinforcers of drug-taking behavior. These findings provide direct evidence for involvement of endogenous cannabinoid systems in brain reward processes. Additional findings include evidence that THC’s reinforcing effects are reduced by an opioid antagonist and reinstated or potentiated by an opioid agonist, and evidence that THC increases β-endorphin in the ventral tegmental area (VTA) and the shell of the nucleus accumbens. Further, augmented levels of β-endorphins in the VTA may be responsible for previous findings of THC-induced dopamine elevations in the nucleus accumbens.
- Presynaptic Effects of Endocannabinoids
Bradley Alger, Ph.D., University of Maryland School of Medicine
Dr. Bradley Alger discussed the discovery of a new mode of cellular communication in which the target (pyramidal) cells may regulate their own state of inhibition by sending a signal backwards across the synaptic junctions (retrograde signaling), causing a temporary inhibition of GABA release. Additionally, Bradley’s team concluded that depolarization-induced suppression of inhibition (DSI) and depolarization-induced suppression of excitation (DSE) are produced by short-term retrograde action of endocannabinoids on presynaptic cells. Further, endocannabinoids inhibit calcium influx, increase potassium efflux, and interfere with the quantal release process. Bradley also noted that prolonged activation of CB1Rs can cause long-term depression of glutamate or GABA release, but that there must be another element to it—it cannot be just a cannabinoid, or must be a different cannabinoid, or it may be that there are different ways in which cannabinoid receptors are activated over a long period of time.
- Efficacy in CB-1 Cannabinoid Receptor Signal Transduction
Allyn C. Howlett, Ph.D., North Carolina Central University
CB-1 receptors exist as a complex with their associated Gi/0 proteins. Association with Gαi1 and Gαi2 occurs via the third intracellular loop and with Gαo and Gαi3, via the juxtamembrane C-terminal domain (fourth intracellular loop). Allyn Howlett detailed evidence showing that structurally different agonists induce conformational changes within these domains, promoting the selective activation of certain G protein subtypes. This may explain the differences in efficacy between WIN55221-2 and the endocannabinoid analog methanandamide. Howlett hypothesized that such selectivity could result in selective G protein-promoted signal transduction pathways.
- Adaptations in a Membrane Enzyme that Terminates Endocannabinoid Signaling
Benjamin Cravatt, Ph.D., The Scripps Research Institute
Dr. Benjamin Cravatt discussed studies involving genetically engineered mice that lack the enzyme fatty acid amide hydrolase (FAAH). The results of these experiments demonstrate that FAAH is a key regulator of anandamide signaling in vivo, setting an endogenous tone that is essential for normal pain transmission. Therefore, Cravatt proposed, FAAH may represent a pharmaceutical target for the treatment of pain and neuropsychiatric disorders. He described his team’s determination of the three-dimensional structure of FAAH by x-ray crystallography, as well as their development of a proteomic assay to screen FAAH inhibitors against whole tissue/cell extracts.
- The Endocannabinoid System and the Regulation of Emotions
Daniele Piomelli, Ph.D., University of California, Irvine
Anandamide is released in selected regions of the brain and is deactivated through a two-step process that consists of transport into cells followed by intracellular hydrolysis. Dr. Daniele Piomelli discussed research that shows that pharmacological blockade of the enzyme fatty acid amide hydrolase (FAAH), which is responsible for intracellular anandamide degradation, produces anxiolytic-like effects in rats without causing the wide spectrum of behavioral responses typical of direct-acting cannabinoid agonists. These findings suggest that anandamide contributes to the regulation of emotion and anxiety and that FAAH might be the target for a novel class of anxiolytic drugs.
Mechanisms of Receptor and Transporter Trafficking
This portion of the program centered on the mechanisms that govern when, why, and how receptor and transporter proteins move within a cell.
- Dopamine Transporter: Are Psychostimulants in Your Neighborhood Forcing You To Move?
Aurelio Galli, Ph.D., Vanderbilt University School of Medicine
Dr. Aurelio Galli presented research in which his team attempted to characterize amphetamine-induced loss of transporter capacity over time. The results showed a temporal correlation between the loss of transporter activity and hDAT cell surface expression. Data suggest strongly that the amphetamine (AMPH) reaction of hDAT transport capacity results from the removal of active carriers in the plasma membrane. Further research implicates DAT trafficking in the hormonal regulation of dopaminergic signaling and shows that insulin is a tool for regulating the AMPH-induced loss of DAT activity and cell surface expression.
- AMPA Receptor Assembly Determined by Q/R Editing
Ingo Greger, Ph.D., Medical Research Council Laboratory of Molecular Biology
Dr. Ingo Greger and his team identified an internal GluR2 pool within the endoplasmic reticulum (ER). Reversion to the exonically encoded Gln resulted in rapid release from the ER and increased surface expression of GluR2. This research leads to the conclusion that Arg607 acts as a master switch, controlling ER-exit. Also, Arg607 may ensure the availability of GluR2 for incorporation into AMPARs. Such a mechanism could explain the presence of GluR2 in the majority of AMPARs in the brain. Greger presented a model in which Arg607 controls receptor trafficking by regulating the assembly of subunit dimmer intermediates into tetrameric channels that are competent for ER export.
- Signaling Mechanisms for Synaptic Plasticity
J. Julius Zhu, Ph.D., University of Virginia School of Medicine
Dr. Julius Zhu described efforts to better understand the signaling that controls AMPA receptor trafficking. His team examined the role of the small GTPases—Ras and Rap—in postsynaptic signaling during synaptic plasticity. Their work showed that Ras relays the NMDA-R and CaMKII signaling that drives the synaptic delivery of AMPA-Rs, with long cytoplasmic tails during long-term potentiation. In contrast, Rap mediates NMDA-R-dependent removal of synaptic AMPA-Rs, with only short cytoplasmic tails that occur during long-term depression. Understanding the signaling pathways that regulate AMPA-R trafficking can help scientists locate potential molecular targets for novel genetic and pharmacological therapies aimed at treating diseases that cause cognitive impairment.
Embryogenesis in Reward-based Behavior
Presentations in this section highlighted several ways in which recent advances in developmental biology can help researchers better understand aspects of prenatal drug exposure and addiction.
- Molecular and Cellular Mechanisms Controlling the Development of Telencephalic Structures
Oscar Marin, Ph.D., University Miguel Hernández
Dr. Oscar Marin described examples of three basic mechanisms that influence the development of brain reward circuitry: patterning, cell migration, and axon guidance. As illustrated by experiments involving genetic manipulation of the transcription factors implicated in their development, differential patterning of the basal telencephalon is likely to generate functional divergence between the shell and core subdivisions of the nucleus accumbens. Alternately, interneuron migration is responsible for increasing the cellular complexity of brain reward circuitry. Finally, axon guidance in the mesolimbic dopamine system is required to effectively wire the brain reward circuitry. Insights into these mechanisms should contribute to understanding normal and abnormal functioning of the brain’s reward system.
- Development of Dopaminergic and Serotonergic Neurons Involved in Reward-associated Behavior in Zebrafish
Su Guo, Ph.D., University of California School of Pharmacy
Reward-associated behavior results when exposure to a substance elicits preferential behavior in favor of obtaining the drug. Dr. Su Guo discussed a series of experiments on drug preference behavior in zebrafish, as well as her team’s use of a zebrafish with mutations that influence the development of dopaminergic and serotonergic neurons in the basal forebrain. From these experiments, her team concluded that zebrafish are an attractive genetic model organism to use in studying the development of specific neuronal types, in this case dopaminergic and serotonergic neurons, as well as the developmental alteration that leads to abnormal reward-associated behavior. This could ultimately allow researchers to link research on genes to neurons, then to circuitries, and ultimately to behavior.
Signal Transduction Mechanisms in Drug Abuse and Addiction
This portion of the program concerned recent discoveries that provide a detailed biochemical characterization of the intracellular signaling cascades involved in drug-induced changes in synaptic transmission and synaptic plasticity.
- GPCR Regulatory Mechanisms: Effects on Tolerance, Sensitization, and Reinforcement
Laura M. Bohn, Ph.D., Duke University Medical Center
Dr. Laura Bohn discussed research aimed at determining if abrogating µOR desensitization affects the reinforcing and psychomotor properties of morphine, leading to the stimulation of the dopaminergic system. Bohn’s team tested morphine and cocaine in assays of locomotor activity, behavioral sensitization, conditioned place preference, and striatal dopamine release in βarrestin2-knockout (βarr2) mice and their wild-type controls. Bohn concluded that GPCR desensitization by GRK and the βarr2 mechanism play an important role in the actions of drugs of abuse. Specifically, the deletion of GRK-6 appears to selectively affect dopamine or dopamine receptors as opposed to opioid receptors, and the deletion of βarr2 appears to select for opiate effects as opposed to only dopaminergic mediated behaviors.
- RGS Protein Function in the Mammalian Brain
Theodore G. Wensel, Ph.D., Baylor College of Medicine
Dr. Theodore Wensel described studies using an RGS9-knockout mice model and a characterization of a retina-specific isoform of RGS9 to examine the molecular pathways regulated by the brain-specific isoform RGS9-2. Wensel explained that his team, along with others, has identified multiple mechanisms for regulating RGS proteins. He concluded that, while the sheer number of such mechanisms is daunting, each of these interactions can be considered a potential site for therapeutic intervention, a discovery that bodes well for addiction research.
- Role of Cdk5 in Drug Addiction
James A. Bibb, Ph.D., University of Texas Southwestern Medical Center
Normal as well as aberrant neurotransmission modulates neuronal circuitry through a complex network of signaling pathways that form the basis of synaptic plasticity. Dr. James Bibb presented evidence that implicates Cdk5 in affecting many of these pathways, including the regulation of protein phosphatases. He concluded that Cdk5 may mediate both functional and structural plasticity and may influence learning and memory. Therefore, Cdk5-dependent changes in neuroplasticity may result in an addicted state.
Neurobiological Basis of Impulsivity
In this section, presenters discussed ways in which impulsivity may play a role in drug abuse.
- Varieties of Impulsivity: Evidence from Animal Studies
John L. Evenden, Ph.D., D.Med.Sci., AstraZeneca
Dr. John Evenden presented a series of drug studies in which rats were used to examine the relationships among the stimulus, response, and reinforcer in the traditional three-term contingency of decisionmaking. The studies revealed that different aspects of impulsivity are differentially sensitive to different drugs and that the effects of drugs depend, in part, on the strain of rat used. Therefore, it can be concluded that impulsive behavior is modulated by a complex interaction of trait factors, such as genetics and personality, and state factors, such as drug treatment. Further, drugs’ effects can vary dramatically if self-control is prompted by negative reinforcement or punishment.
- Behavioral Models of Impulsivity in Humans and Nonhumans: Effects of Drugs
Harriet de Wit, Ph.D., University of Chicago
Dr. Harriet de Wit discussed studies implying that there are at least two separate processes underlying impulsive behaviors – impairments in decision making and planning and impairments in behavioral inhibition. Her team used human and nonhuman laboratory models to investigate these processes and concluded that, while delay discounting procedures and the stop task are both valid methods for measuring impulsive behavior, delay discounting and behavioral inhibition reflect separate processes. De Wit stated that these findings are a starting point for identifying both the underlying factor structure and the neurobiology of impulsive behavior.
F. Gerald Moeller, M.D., University of Texas Health Sciences Center
Dr. Gerald Moeller presented a review of the literature and new data on the associations among psychostimulant abuse, impulsivity, and brain function, which show that impulsivity appears to be a risk factor for stimulant abuse. Further, impulsivity is associated with changes in brain function, and these changes are potentially related to the chronic effects of stimulants. This information provides a foundation for research that will investigate potential treatments for impulsivity and associated stimulant abuse.
William G. Iacono, Ph.D., University of Minnesota
Dr. William Iacono discussed the relationships among externalizing psychopathologies that are indicative of behavioral disinhibition. He described a study in which P3 amplitude elicited from a visual oddball task in a population-based sample of a group of 17-year-old males and females. Reduced P3 amplitude in males was associated with the presence of ADHD, conduct disorder, and substance use disorders, as well as with the presence of substance use disorders in their parents. For females, reduced P3 amplitude was associated with having a substance use disorder only. These findings suggest that a dimension of disinhibitory psychopathology indexed by reduced P3 amplitude underlies the relationship among childhood externalizing and substance use disorders, especially in males.
Neurobiological Mechanisms of Drug and "Natural" Reward
In this segment, researchers focused on identifying the neurobiological substrates involved in learning, motivation, and memory with regard to natural rewards.
- The CNS Regulation of Food Intake: Peripheral Signals and CNS Effector
Randy J. Seeley, Ph.D., University of Cincinnati College of Medicine
Dr. Randy Seeley of the University of Cincinnati College of Medicine described research analyzing neurotransmitters that have been linked to food intake in order to discern which of the transmitters are more important than others in regulating food intake. The precise location of the opioid receptors that are important in influencing the activity of these systems is unknown. Also unknown is the relationship of the classic reward circuits involved in the dopamine system and their ability to interact with various peptide systems. These are critical issues in terms of understanding natural rewards, and of understanding why the dietary environment that we live in leads us to the propensity to gain weight.
Jaak Panksepp, Ph.D., Bowling Green State University
Dr. Jaak Panksepp discussed his theory that, in order to understand drug addiction, researchers must understand the nature of affective processes. Further, affective states of the nervous system are emergent evolutionary properties of complex neural dynamics that should be conceptualized properly and linked to the concrete, multidimensional brain processes from which they emerge. Panksepp summarized a variety of research on affective processes and concluded that an understanding of these processes may be necessary to understanding addictions.
- The Expression of a Simple Appetitive Response Becomes Dopamine-Independent with Overlearning
Jon Horvitz, Ph.D., Boston College
Dr. Jon Horvitz described a series of head-entry experiments that showed that once a reward-directed behavior becomes habit, the expression of the response seems to become dopamine independent. Further, dopamine is needed to execute internally-generated responses, but not to respond to an overlearned conditioned stimulus. Horvitz noted that when a person goes to a clinic for treatment, dopamine antagonist drugs are surprisingly ineffective at reducing drug-seeking behavior. Horvitz proposed that by the time a person goes to a clinic, the drug-seeking behavior has been repeated so many times and has been tied so strongly to eliciting stimulae that the behavior actually becomes independent of dopamine transmission.
Structure, Function, and Regulation of the Dopamine Transporter
Presentations in this section addressed ways in which the dopamine transporter may function as a multimer or an ion channel, in association with regulatory proteins and human disease.
- Oligomerization of the Dopamine Transporter: Cocaine-Induced Conformational Changes at a Homo-Dimer Interface
Jonathan A. Javitch, M.D., Ph.D., Columbia University
Dr. Jonathan Javitch discussed evidence supporting the idea that the dopamine transporter (DAT) and homologous neurotransmitter exist in the plasma membrane as oligomers. Cysteine cross-linking experiments in DAT show that TM6 and TM4 contribute to the formation of two distinct symmetrical interfaces and suggest that the transporter exists as a tetramer. Cocaine analogs prevent cross linking of some, but not all, cysteines in TM4, which suggests that cocaine binding induces a conformational change at the multimer interface.
- Protein-Protein Interactions: Defining New Pathways Involved in the Regulation of the Dopamine Transporter
Gonzalo E. Torres, Ph.D., Duke University Medical Center
Dr. Gonzalo Torres discussed the identification of protein-protein interactions supporting dopamine transporter trafficking and its result in increased understanding of mechanisms that underlie the transporter’s cellular regulation and functional expression. Torres discussed attempts to use a proteomic approach to identify the entire network of proteins associated with DAT through selective immunoprecipitation of the transporter complex from examining brain tissue, separating the proteins using 2D gels, and analyzing the proteins using mass spectrometry and database analysis. Future research will focus on identifying the protein network associated with monamine transporters, as well as the subcellular compartments involved in the different steps of targeting and trafficking these transporters, with the ultimate goal of examining how these mechanisms relate to the problem of drug addiction.
- DA Transporter Currents: A Reason for Excitement
Susan L. Ingram, Ph.D., Washington State University
Dr. Susan Ingram described research examining the concept that DAT may dynamically regulate DA neurotransmission through both uptake and the release of DA through its channel activity. She discussed research indicating that DAT mediates an uncoupled chloride conductance and that DAT currents in neurons are activated at much lower concentrations than the D-2 receptor or transport. Other studies showed that the DAT-mediated chloride conductance excites DA neurons, and that activation of DAT-mediated currents is a potential mechanism for stimulating DA release in the substantia nigra.
- Dopamine Transporter Genetics: From Model Systems to Man
Randy D. Blakely, Ph.D., Vanderbilt University School of Medicine
Dr. Randy Blakely highlighted the ways in which efforts to manipulate the DAT of C. elegans using a forward genetic approach have elucidated critical sites supporting DAT function and have advanced a new model system for the study of dopamine neuron degeneration. Included was the finding that non-DAT-1 genes participate in DAT-1 function, DAT-1 regulation, or in the sensitivity to neurotoxin. Additionally, he discussed human studies on the elucidation of DAT-supported human disorders via functional investigations of DAT polymorphisms.