Charles Ouimet - US grants

The long term goal of this research is to increase our understanding of plasticity in neurons postsynaptic to dopamine- terminals in limbic or paralimbic brain regions. Dopaminergic mechanisms and limbic and paralimbic brain regions are related to the etiology of schizophrenia and affective disorders. The anterior cingulate cortex is a paralimbic region that is dopamine- enriched and forms an interface between the DARPP-32 and ARPP-21, in dopaminocptive neurons in anterior cingulate cortex, is suggested in this proposal. DARPP-32 and ARPP-21 are enriched in dopaminoceptive cells containing the D-1 dopamine receptor, and will be studied with neuroanatomical and neurochemical methods. DARPP-32 is regulated by dopamine and it is very likely that ARPP- 21 is also regulated by that transmitter agent. Both phosphoprotein are present in anterior cingulate cortex; ARPP-21 is enriched in neurons in the upper layers of carted that receive the superficial plexus of dopaminergic axon terminals and DARpp-32 is enriched in neurons in the deep layers that receive the deep plexus of dopaminergic axon terminals. DARPP-32 acts as a "third messenger" for dopaminergic activation of the dopamine D-1 receptor (dopamine is the first messenger and cyclic AMP is the second messenger), and these is very likely true for ARPP-21 as well. The goals of the specific aims of this proposal are to define the morphology of DARPP-32- and ARPP-21- containing neurons in the singulate carte of untreated ("normal") rats, to determine whether or not these neurons receive classical dopaminergic synapses, to determine whether decreased dopaminergic input can induce a compensatory increase in the synthesis os mRNA for DARPP-32 and ARPP-21, and to assess the effects of decreased dopaminergic input on the "Golgi-morphology" of the DAEPP-32- and ARPP-21-containing neurons. Changes in the Golgi-morphology of these cells many also reflect an attempt to re-establish homeostasis. The techniques employed will include light and electron microscopic immunocytochemistry, double staining immunocytochemistry, combined Golgi staining and immunocytochemistry, in situ hybridization, and biochemical analysis with Western blotting. The study of dopamine and dopamine-regulated phosphoprotein anterior cingulate cortex should increase our knowledge of dopaminergic function in cortex and demonstrate whether these two phosphoprotein, and the cells that contain them, can be altered by decrease dopaminergic activity.

DESCRIPTION (Adapted from applicants abstract): The long term objective of this research is to identify the mechanism(s) involved in the etiology of depression and its relief by antidepressants. Antidepressants can be broadly classified into three types: Those that inhibit monoamine oxidase, those that block re-uptake of monoamines and those that specifically block re-uptake of serotonin. The fact that these antidepressants work on different mechanisms suggests that the biochemical cascades they initiate ultimately converge on a common process. Is altered neuronal circuitry, indicated by significant and selective changes in dendritic spine density, a part of that common process? The specific aim of this proposal is to use a novel method for estimating dendritic spine density to test the hypothesis that chronic exposure to the 3 different types of antidepressants causes specific circuits in the brain to become rewired. Histochemistry for two novel markers for dendritic spines (spinophilin and phallodin) will be used in conjunction with confocal laser microscopy, a novel modification of the "dissector" technique, and a novel computerized imaging and quantification system. The proposed studies should: 1) demonstrate that the new method has merit; and 2) directly test the hypothesis that antidepressants cause significant changes in brain wiring as evidenced by altered dendritic spine densities. The new technique should be broadly applicable to other studies on brain plasticity. A finding that antidepressants cause specific parts of the brain to be rewired would identify a new mechanism by which antidepressants may work and serve as a building block on which many new studies investigating depression and its treatment could be based.