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.
While neuronal activity is generally on the timescale of tens of milliseconds, many high-order brain functions require evidence that accumulates over many seconds minutes or hours. In this proposal, I develop a new system and new tools for understanding the molecular and circuit bases of interval time as it relates to motivation. The findings from these studies will suggest new targets and strategies for intervention in the many disorders of motivation and other long-lasting brain functions.