Dr. Crowder proposes to identify and characterize genes that control volatile anesthetic action in the nematode Caenorhabditis elegans in the hope that such findings will eventually be useful in the design of better anesthetics and in defining mechanisms responsible for anesthetic-induced nervous system depression. In preliminary studies, recombinant inbred strains of C. elegans have been found variant in their sensitivity to halothane, with halothane EC50 concentrations of 0.2 - 0.9 percent atm being in the same range of concentrations administered during surgical anesthesia. Genetic mapping of the halothane sensitivity phenotype in these recombinant inbred strains has identified 5 genetic loci that influence halothane potency. Dr. Crowder proposes to 1) continue identifying and mapping in recombinant inbred strains the loci that control volatile anesthetic action in C. elegans, which are presumed to code for targets or modulators of anesthetic action; 2) map individual loci in congenic strains, which allow determination of the independent effect of each locus on anesthetic sensitivity; 3) positionally clone the strongest loci controlling anesthetic sensitivity in C. elegans, which involves injecting the cloned DNA of the locus associated with anesthetic sensitivity into the gonad of an anesthetic-resistant animal and screening the progeny for the anesthetic-sensitive phenotype; 4) analyze the sequence of genes that control anesthetic sensitivity; and 5) characterize the expression of cloned genes, including RNA expression and the distribution of the actual gene product. It is hoped that these results will help guide in vitro and in vivo studies in higher organisms.
