A major event following brain trauma is the loss of autoregulatory capacity of brain microvessels which results in sustained hypoperfusion and improper delivery of vital metabolites to brain tissue. Previously we established that the sustained hypoperfusion and the lack of autoregulatory capacity post trauma are largely due to the capability of injured cells in the vascular wall and in other brain compartments to synthesize and maintain excessive amounts of endothelin-1 (ET-1). ET-1, as well as its two isoforms ET-2 and ET-3, exert signal-transducing, vasoconstrictor effects through their only known receptors ET-RA and ET-RB. Although activation of both receptors is known to cause vasoconstriction in both in vitro preparations and non-neural tissues, little is known of their role in the injured brain and the development of secondary cell injury. In fact the role of ET-RA and ET-RB in vasoconstriction and their pharmacological manipulation to prevent cell injury after traumatic brain injury (TBI) have never been studied. ? ? We propose that upregulation of the genes encoding for the synthesis of ET-RA and ET-RB in smooth muscle and endothelium (where they are expressed most abundantly), as well as in other perivascular cells, underlie the enhanced contractility previously observed in the microvascular wall after TBt. In this proposal our specific aims are to: 1) characterize quantitatively in time the expressions of ET- RA and ET- RB (mRNA) in distinct cellular compartments of the vascular wall and surrounding neuropil prior to and after TBI, 2) translationaly inhibit the expression of ET- RA and ET- RB before TBI to improve the impaired microcirculation, and 3) pharmacologically attenuate hypoperfusion post TBI by application of selective antagonists to ET- RA and ET- RB. ? ? We will use in situ hybridization to measure mRNA synthesis and in vivo intracerebroventricular application of antisense oligodeoxynucleotides to attenuate ET- RA and ET- RB gene expressions."""""""" In tandem use of selective receptor antagonist with the 14C iodoantipyrine technique we will assess antagonist effects on cerebral cortical blood flow. By similar in tandem use of antisense application and the 14C iodoantipyrine technique we will determine the reciprocal effects of the receptors and their antgonist on each other and respective affect on the microcirculation after TBI. Histological and motor behavior studies will be used to assess neurological outcome following the therapeutic interventions. ? ? The results will provide valuable information on the expression of endothelin receptors (mRNA and proteins), their cellular distribution and the causal relationships of this expression to the alterations in the brain microcirculation after TBI. Precise times of ET- RA and ET- RB expressions will be established that should allow us to establish effective therapeutic interventions. The study will also provide direct evidence on the therapeutic value of suppressing gene regulation or receptor activation on cortical perfusion, histologic or neurologic injury, and its effect on impaired motor skills after TBI. ?
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