Worm Breeder's Gazette 11(4): 106
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
A series of three muscle contractions occurs in C. elegans during defecation (Jim Thomas, Genetics 124:855-872, 1990): (1) the posterior body muscle contraction (pboc) forces the gut contents forward, (2) the anterior body muscle contraction (aboc) forces the gut contents backward, and (3) the expulsion muscle contraction (exp) forces the gut contents through the anus. This sequence is repeated every 50 seconds. Two GABAergic neurons, AVL and DVB, are jointly required for exps. As a first step toward identifying the neurons involved in defecation, we have shown that unc-25 animals, which lack immunoreactivity to the neurotransmitter GABA, are constipated due to a reduction of exps. Killing two GABAergic neurons, AVL and DVB, nearly eliminates exps producing a constipated phenotype very similar to that of unc-25. Killing either DVB or AVL alone decreases the frequency of exps, but does not lead to constipation. In addition, AVL is also required for abocs. Laser ablation of AVL results in animals with only very rare abocs. Thus, this neuron controls two of the muscle groups that contract during defecation. This result is more intriguing because unc-25 animals, which lack detectable GABA staining, have normal abocs. Based on these results, we propose that AVL stimulates abocs via a second neurotransmitter. Interestingly, Jim Thomas has identified six genes (denoted aex genes) that when mutated cause constipation due to defects in both abocs and exps. Perhaps these genes encode molecules required for the development or function of AVL and DVB, or of cells that mediate the effects of AVL and DVB. Using the known connectivities of AVL and DVB, we are currently expanding our laser experiments to identify other neurons that control defecation movements. Existing mutations that eliminate the function of AVL or DVB have been identified using laser surgery. The above experiments suggest a strategy for identifying mutations that affect the function of either AVL or DVB. Mutants lacking AVL should become constipated after killing DVB, while mutants lacking DVB should become constipated after killing AVL. We tested these predictions using mutants we had previously shown to lack AVL (unc-73) or DVB (sem-4). In 70% of unc- 73(e936) animals the AVL axonal process, which is normally found in the ventral nerve cord, is found in the dorsal cord. As a consequence, the average frequency of exps in unc-73 animals is reduced to 50%; however, unc-73 animals are not constipated. In 70% of unc-73 animals, killing DVB reduces the frequency of exps to less than 10% and results in severely constipated animals. In 50% of sem-4(n1378) animals DVB is missing as assayed by GABA expression (M. Basson, personal communication). sem-4 animals miss half of their expulsions, but are not constipated. In 50% of sem-4 animals, killing AVL reduces expulsions to less than 10% and causes the animals to become constipated. We have begun to screen through known mutants using the laser to identify those lacking AVL or DVB function. So far we have found that unc-2, lack AVL function. We are conducting genetic screens for new mutations that eliminate the function of AVL and DVB. Among mutations that make unc-73 or sem- 4 animals constipated should be mutations that specifically disrupt DVB and AVL function, respectively. In addition to these synthetic- constipated mutants, such screens should also generate mutants that are defective in other steps of defecation, similar to those first described by Jim Thomas. We have screened 7,000 unc-73 genomes and are presently characterizing 100 constipated strains. Similarly, we have screened 10,000 sem-4 genomes and are characterizing 34 constipated strains.