Mutations in Rho GTPase effectors and regulators cause mental retardation in humans. It has been assumed that these defects are caused by developmental defects given the important role of Rho GTPases in regulating the cytoskeleton and thus cell migration. However, in at least one case, the RhoA GAP oligophrenin, restoration of gene function in adult neurons restores at least some of the observed synaptic defects. This suggests an important role for RhoA in control of adult neuronal activity. Previously we have demonstrated a role for the single C. elegans, RhoA ortholog, RHO-1, in control of actylcholine (ACh) release from cholinergic motorneurons. RHO-1 acts as part of a network of G alpha signaling pathways that regulate neuronal activity by regulating both production and destruction of the second messenger diacylglycerol (DAG), which is a regulator of synaptic vesicle release. RHO-1 can be activated by at least two G alpha pathways, resulting in increased ACh release. GPA-12 (Ga12) acts via the RHGF-1 RhoGEF which activates RHO-1. EGL-30 (Gaq) acts via the UNC-73 RhoGEF which activates RHO-1. Hyperactivation of RHO-1 in cholinergic motorneurons causes hypersensitivity to the acetylcholinesterase inhibitor aldicarb suggesting elevated levels of ACh release, whereas inhibition of RHO-1 results in resistance to aldicarb suggesting decreased ACh release. However,
rhgf-1 mutants have no phenotype in the absence of activated GPA-12 and UNC-73 mutants, although very lethargic, have a normal response to aldicarb. We tested whether these two RhoGEFs were redundant by constructing an
unc-73;
rhgf-1 double mutant. However, these mutants also had a wildtype response to aldicarb. Thus, either another RhoGEF acts to regulate ACh release, or a decrease in ACh release in
unc-73 mutants is compensated by an increase in muscle response to ACh. RhoGTPases are activated by RhoGEFs and inhibited by RhoGAPs. To identify the RhoGAPs that regulate RHO-1 function in adult neurons we tested viable RhoGAPs for response to aldicarb. Two mutants were both hypersensitive to aldicarb,
rga-3 and
rga-4. RGA-3 and -4 are RhoGAPs that have redundant roles in regulating C. elegan embryogenesis. Experiments are ongoing to test if
rga-3 and
rga-4 act within cholinergic motorneurons to negatively regulate RHO-1 and thus decrease levels of ACh release.