The objective of this proposal is to examine the relationship of the amphetamine (AMPH)-induced increase in striatal Ca2+-binding protein, calmodulin (CaM) to the expression of sensitization to AMPH. Repeated, intermittent treatment of humans and animals with AMPH leads to a lasting dopamine (DA)-related behavioral sensitization, paralleled by enhanced stimulus-induced DA release in rat striatum and nucleus accumbens. Repeated AMPH elicits an enduring withdrawal-dependent increase in rat striatal and limbic forebrain CaM. This proposal will test the hypothesis that an alteration of the content and/or functional activity of CaM parallels development and contributes to expression of AMPH-induced sensitization. In particular, I hypothesize that an increased CaM Content or phosphorylating activity in striatal synaptosomes contributes to the enhanced DA release in AMPH-sensitized rats through increased phosphorylation of synapsin I and neuromodulin.
The specific aims of this proposal are to: (I) Relate the increase in CaM to AMPH-induced sensitization by determining whether the AMPH-induced increase in CaM can be blocked by drug treatments (haloperidol, MK-801) known to block the development of behavioral sensitization to AMPH. (2) Determine whether acute and/or repeated AMPH elicits changes in endogenous phosphorylation of synapsin I in rat striatum using a phosphosynapsin I antibody directed against a CaM-Kinase II phosphorylated site. Parallel changes in activities or content of protein kinase C, CaM-Kinase II and protein phosphatases will be determined. (3) Investigate the ability of AMPH to enhance phosphorylation of neuromodulin and synapsin I and alter kinase and phosphatase activities in a synaptosomal preparation. The effect of various stimuli (AMPH, K+, 4-AP) on DA release in synaptosomes from AMPH-pretreated rats will be examined. (4) Investigate an involvement of postsynaptic CaM in sensitization by measuring changes in CaM content, CaM-binding proteins, CaM-Kinase II activity and content and PKC activity in three postsynaptic membrane preparations in striata from rats pretreated with saline or AMPH: a) sucrose-density purified postsynaptic membranes; b) purified postsynaptic densities and c) total and postsynaptosomal cytoskeletal fractions. AMPH-induced sensitization exhibits many characteristics possessed by other models of synaptic plasticity in which Ca2+ and CaM play a definite role. The fact that the increase in CaM exhibits characteristics that parallel aspects of behavioral sensitization, such as anatomical specificity, withdrawal- dependence and endurance, suggests a high demand for a Ca2+- and CaM- stimulated process in sensitization.
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