BRAIN Initiative K99 Project

National Institute On Alcohol Abuse And Alcoholism

Investigators

Linked publications & trials

Abstract

In accordance of aims of the proposal, the following experiments were done to determine the role of dopamine D2 receptor (D2R) containing neurons in the nucleus accumbens (NAc) on food intake: 1- The NAc is the ventral region of the striatum, which is a brain region implicated in reward-related behavior. In a preliminary study, Adora2a-Cre mice were injected with the Cre-dependent excitatory DREADD, hM3Dq, targeted at the nucleus NAc. This was done to specifically deliver the excitatory DREADD to the D2R-containing projection neurons in this brain region, which we could then activate using the synthetic agonist clozapine-N-oxide (CNO). Following viral introduction, mice were singly housed in specialized home cages, that continuously monitor intake of standard rodent chow and water, as well as locomotion. After four weeks of viral transfection, mice were injected with saline (5 mL/kg) for three consecutive days to habituate them to handling and injections. The final day of saline injection was used to determine baseline intake and activity, to allow for within-subjects analysis. On the fourth day mice received CNO (3 mg/kg, i.p.) and food and water intake, as well as locomotion, were recorded continuously for 24 h. Consistent with the hypothesis, activation of the NAc D2R containing neurons with peripheral administration of CNO seems to drive intake of a standard rodent chow. 2- This preliminary study relied on the activation of all D2R-containing projection neurons in the NAc, but is crucial in that it provides motivation to further explore specific subpopulations of these neurons that might be important in driving food intake. Specifically, we are examining the effects of the NAc D2R-containing neurons that project to the ventral pallidum. This is being done by injecting the retrograde CRe-dependent hM3Dq into the ventral pallidum of Adora2a-Cre mice. This will result in DREADD localization specifically in the D2R-containing neurons projecting from the NAc to the ventral pallidum, as described in my original proposal to selectively decrease the expression of D2Rs in the projection neurons of the striatum, including the NAc. D2 receptors are Gi/o-coupled receptors, which exert inhibitory control over a number of neurochemically distinct cell populations within the striatum. Therefore, removing D2Rs from the projection neurons eliminates the inhibitory actions of those receptors; this is akin to removing the break and allows the GABAergic projection neurons to more easily fire. When I selectively knocked down the D2 receptors from NAc projection neurons the mice consumed more standard rodent chow compared to littermate controls, and showed subsequent weight gain. 3. In order to further determine how the brain, and striatum specifically, might be interacting with the peripheral hunger and satiety signals, I turned my attention to insulin, a peptide hormone produced in the pancreatic beta cells. In the periphery, insulin is known to regulate metabolism by promoting the absorption of glucose from the blood. I noticed that my mice with low levels of D2 receptors on the striatal projection neurons had low fasting insulin levels compared to littermate controls (Figure 3A). Insulin produced in the periphery crosses the blood-brain barrier, and has known actions in several brain regions, including the striatum. Using qPCR techniques, I determined that mice with low levels of D2 receptors on the striatal projection neurons had increased insulin receptors in the same region. Further, this increase in striatal insulin receptors had profound effects on dopamine release. Using slice fast-scan cyclic voltammetry I replicated findings that insulin exposure leads to an increase in striatal dopamine release and found that this is amplified in mice lacking D2 receptors on the striatal projection neurons (Figure 3C). While there is still much to do, these findings begin to provide a mechanistic understanding of how the striatal circuitry might be integrating peripheral signals in the control of food ntake.