Serotonin increases the rate of pharyngeal pumping and egg laying, decreases foraging behavior and is required for proper male mating behavior. Serotonin can influence pharyngeal pumping rate by stimulating MC, a motor neuron which stimulates pharyngeal muscle contraction, or by acting directly on pharyngeal muscle. To determine the receptor or receptors that mediate this response, we have searched the genomic sequence for candidate genes having sequence identity to known vertebrate and invertebrate serotonin receptors. We found seven candidate genes using this method: C02D4.2, C09B7.1, C52B11.3, F01E11.5, F14D12.6, F59C12.2 and M03F4.3. Two of these genes (F59C12.2 and M03F4.3) were previously identified as possible serotonin receptor candidates (Ribeiro and Hamdan IWM97, p278
.) We were not able to distinguish octopamine receptors from metabotropic serotonin receptors based on sequence. In addition, there are no sequences in the C. elegans database with significant similarity to the ionotropic serotonin receptor subclass, 5-HT3. We have created GFP fusion constructs of four of the candidate genes (C02D4.2, F01E11.5, F14D12.6 and F59C12.2) using overlap extension PCR. We directly injected these PCR products into worms to determine GFP expression patterns. F01E11.5 and F14D12.6 show limited expression outside the pharynx. The F01E11.5 construct expresses in a few unidentified neurons of the head, while F14D12.6 expression is most pronounced in the spermatheca of the adult. Two of the genes, C02D4.2 and F59C12.2, show pharyngeal expression of the fusion construct. F59C12.2 shows nearly ubiquitous pharyngeal and extrapharyngeal staining. C02D4.2 pharyngeal muscle expression is specific for the metastomal flap muscles, pm1
, and the anterior terminal bulb muscles, pm6
. The C02D4.2 construct also expresses in muscles of the head, the pharyngeal serotonergic neuron NSM, the ventral nerve cord and unidentified neurons in the nerve ring, head and tail regions. In males, C02D4.2 expresses in dorsal and ventral body muscles of the tail region. The construct also expresses in the male-specific diagonal muscles and serotonergic CP neurons. The expression pattern of C02D4.2 appears to be consistent with several known effects of serotonin. We are currently working to knock out both C02D4.2 and F59C12.2 using PCR detection of EMS-induced deletions of these genes.