Behavioral Functions of Neuropeptides

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Our laboratory is engaged in investigating the behavioral concomitants of the inhibitory effects of galanin. The neuropeptide galanin is localized in the hippocampus, coexists with acetylcholine in the rat septohippocampal pathway, coexists with norepinephrine in the locus coeruleus, inhibits the release of glutamate, acetylcholine, serotonin, and norepinephrine, and inhibits evoked adenylate cyclase signal transduction. Galanin is overexpressed in the basal forebrain in Alzheimer's disease. Our past experiments revealed that central microinjection of galanin in rats impairs performance on several learning and memory tasks. We discovered that galanin overexpressing transgenic mice (GAL-tg) display analogous deficits on learning and memory tasks including the Morris water maze spatial learning probe trial, olfactory memory in social transmission of food preference, and trace cued fear conditioning, when compared to WT littermate controls. GAL-tg mice represent a mouse model of galanin overexpression in Alzheimer's disease, which can be used as a translational tool to test hypotheses and generate novel therapeutics for treating the memory loss that characterizes Alzheimer's disease. This year postdoctoral fellow Nathan Rustay sought to increase the neuroanatomical specificity in our modeling of galanin overexpression in mice. Dr. Rustay employed an adeno-associated viral vector (AAV) containing the galanin gene, generated by Professor Thomas McCown at the University of North Carolina and kindly contributed to our laboratory. Methods followed Dr. McCown's protocol, which had successfully demonstrated anti-seizure activity with his AAV-galanin. Collaborator Dr. Markus Heilig and members of his NIAAA laboratory kindly contributed expertise in training Dr. Rustay on sectioning, immunocytochemical visualization of the green fluorescent protein (GFP) marker, and in situ hybridization for galanin mRNA. Dr. Miles Herkenham, NIMH, kindly permitted Dr. Rustay to section the mouse brains on his cryostat in our PNRC Pod 1C. First experiments included microinjections of the AAV-galanin into the medial septum-diagonal band or into the ventral hippocampus. Neuroanatomical analyses demonstrated the presence of GFP and of the galanin gene at both sites. Behavioral analyses were conducted across a time course of 2 weeks to 2 months. Two learning and memory tasks, Morris water maze spatial learning and trace fear conditioning, evaluated cognitive function. AAV-galanin treated C57BL/6J mice displayed no significant differences in performance on these tasks, as compared to AAV-GFP controls. These findings indicate the need for more global galanin overexpression, at high concentrations in multiple brain regions, to produce memory impairments. Results are consistent with clinical findings that widespread galanin overexpression coincides temporally with mid- to late-stage Alzheimer's disease, when cognitive functions are significantly compromised. This year postdoctoral fellow Kathleen Bailey expanded on the first full behavioral phenotyping characterization of a new galanin subtype receptor GAL-R2 knockout mouse. The mutant line was generated by collaborator Dr. John Hohmann at Nura, Inc. in Seattle. GAL-R2 knockout mice were normal on measures of general health, home cage behaviors, neurological reflexes, sensory abilities, and motor functions. Dr. Bailey had shown that GAL-R2 null mutants did not display deficits on trace fear conditioning or on spatial navigation in the Morris water maze. This year Dr. Bailey confirmed and extended her initial findings of an anxiogenic-like phenotype in GAL-R2 null mutants on the elevated plus maze. Phenotypes on other measures of stress and anxiety-like traits, including stress-induced hyperthermia and light/dark exploration, were similar across gentoypes, indicating a highly selective anxiety-like effect of the GAL-R2 mutation in the elevated plus maze. Further, Dr. Bailey's experiments with GAL-R2 mice were extended to test social interactions and anxiety-like behaviors in a social setting. No genotype differences were detected in our automated 3-chambered social task. Dr. Bailey's findings support the emerging evidence that galanin is an inhibitory neuromodulator with anxiolytic actions in specific conflict situations.