Michael A. Crickmore, Ph.D. - US grants

Summary: It is well known that individuals differ in drug tolerance and propensity for addiction, and that both of these traits are highly heritable, resulting from interactions of many alleles. This multi-genic heritability makes diagnosing susceptibility and targeted treatment more difficult than for monogenic disorders. Despite the vast genetic heterogeneity, there are core principles of abuse and addiction that provide possibilities for new treatments: all drugs of abuse derive their hedonic qualities by upregulating dopamine signaling, and alterations in dopaminergic circuitry following repeated exposure are the root cause of addiction. It is therefore likely that many of the relevant genetic susceptibility loci are involved in the regulation of dopamine responses to drugs and triggering situations. If we refine our ability to precisely manipulate dopaminergic circuitry, we will be able to design treatments to counter the causes of abuse and addiction that are tailored to the individual, without necessarily needing to correct the causal alleles. The importance and potential of studying dopaminergic control systems to combat addiction have long been recognized; I propose a new and promising approach to rapidly identify the circuit and molecular principles of dopaminergic regulation. My lab has recently established two new systems for studying dopaminergic control of motivated behavior. In these systems, two separate populations of dopaminergic neurons provide dynamic motivational input into two distinct aspects of male mating behavior in Drosophila melanogaster: courtship and copulation. The small populations of dopaminergic neurons that control these behaviors are embedded within neural networks that precisely tune the amount of dopamine released so that the level of motivation matches the relevance of the behavioral goals. We study these behaviors because i) they show clear hallmarks of dopaminergic regulation of motivation; ii) they are robust, unambiguous, and easily quantified; iii) the underlying neurons are identifiable and genetically accessible through their expression of sexually dimorphic genes; and iv) the history of work in Drosophila suggests that the principles we uncover will apply to mammals. The main goal of this work is to generate new hypotheses and drug targets for interventions that will prevent the onset and persistence of drug addiction. The characterization of novel regulators of motivational dopaminergic circuitry will also be of use in identifying people at high-risk for abuse and addiction through their possession of altered alleles at these loci. This project is innovative because it combines circuit and molecular approaches in a simple model system to rapidly identify behaviorally-relevant regulators of dopaminergic activity. I do not believe that any such approach has been taken before and it therefore promises new discoveries and potential for treatments and diagnostics.

Project Summary: The decision to commence a new behavior often requires termination of the ongoing behavior. This implies that the many drive states produced by an animal impact not only the neural circuits underlying their directly associated behaviors, but also those of many other behaviors. My lab has shown that the mating behaviors of male Drosophila are under motivational control and may be abandoned in the presence of stimuli evoking competing drives?depending on the relative intensities of the contending drives. These behaviors are motivated by dopaminergic neurons, one of many features shared with human motivational control. I present preliminary data demonstrating that multiple competing drives integrate synergistically to cause male Drosophila to prematurely terminate copulations that would last ~23 minutes if left undisturbed. This integration occurs at a set of eight male-specific, GABAergic Copulation Demotivating Neurons (CDNs) that cause immediate termination when stimulated beyond a threshold and can integrate diverse inputs over long timescales. During the first 5 minutes of mating, even the most severe threats cannot stimulate the CDNs and therefore do not cause termination; but as the mating progresses the CDNs become more accessible to diverse demotivating stimuli, gradually permitting termination in response to weaker and weaker inputs. I present a computational model for synergistic integration of competing inputs at behavior-specific demotivating neurons, demonstrating how this circuit logic can promote either behavioral stability, or flexibility, depending on the individual strengths of the full complement of drive states. I also propose a novel hypothesis for how behavior- specific demotivating neurons increase their sensitivity as the goals of the behavior are achieved. These experimental findings place the rarely studied topic of demotivation at the center of behavioral decision making and our computational work suggests several novel and testable hypotheses. The main goals of this grant are i) to understand how information from competing drives is processed and delivered to behavior-specific demotivation circuitry; and ii) to understand how this information is integrated with the motivational state of the ongoing behavior to decide whether or not to switch behaviors. This work will establish a new, front-line model system for high-order interactions between multiple motivations, with strong indications that the principles and models we derive will provide a framework for understanding motivations and decision making in humans. Project Relevance: This project explores a fundamental but understudied principle of motivational regulation: demotivation as goals are achieved and circumstances change. Dysregulation of motivation is central to drug addiction, depression, compulsive disorders, among many other behavior and mood disorders. The robust behaviors and precise manipulations in this proposal will relate neuronal activity to behavior and allow a direct attribution of causality. The data collected will be used to extend our circuit and computational models that are generalizable across animals and behaviors.

Project Summary Understanding how neuronal networks construct long lasting and slowly evolving states is an outstanding problem in behavioral neuroscience, both basic and disease-related. My lab focuses on motivation, as its dysregulation is central to addiction and mood disorders. Motivations evolve over minutes to hours, much longer than the timescale of standard neuronal processes, with membrane capacitive time constants of 10-100 milliseconds. The circadian clock keeps intracellular time through transcriptional and translational oscillators, but this mechanism is likely too slow to accurately measure the shorter time periods relevant for most behaviors. We have recently developed mating duration in Drosophila as a powerful system for exploring a change in motivation over time as behavioral goals are achieved. At six minutes into the mating, sperm is transferred from the male to the female and a dramatic shift takes place within the male's nervous system: he will no longer sacrifice his life to sustain the mating. These simultaneous events are caused by the output of four male-specific neurons that produce the neuropeptide Corazonin (Crz). If the Crz neurons are inhibited sperm is not transferred and the male does not downregulate his motivation, leading to matings that last for hours instead of the usual ~23 minutes. We exploit the robustness, experimental tractability, and neuronal localization of these phenomena to gain insights into the molecular and circuit bases of interval timing. Our preliminary data point to CaMKII as a molecular interval timer that functions to delay the activity of the Crz neurons for the first six minutes of mating. The timer works through the gradual decay of sustained autophosphorylation following an initial rise in calcium. This proposal centers on understanding i) how the decay rate of CaMKII is tuned to measure out various time intervals in different neurons, and ii) how the CaMKII timer is read out and translated into a timed signal. We have identified multiple candidate factors that may work to sculpt the rise and fall of CaMKII activity, and thereby set the time interval to be measured. For the timing mechanism itself, I propose to test the hypothesis that CaMKII activation prevents the accumulation of cyclic AMP that would otherwise arise from mutual excitation within the Crz network during mating. The decay of CaMKII activity allows super-threshold cyclic AMP accumulation, leading to a large calcium influx that synchronizes the four Crz neurons and generates a single event that drives sperm transfer and the shifts the motivational state at six minutes after the initiation of mating. This would be the first mechanistic description of a neuronal interval timing system. The high conservation and ubiquitous expression of the molecules involved suggest similar functions in long-lasting brain functions across the animal kingdom, including humans.