Serotonin (5-HT) stimulates both pharyngeal pumping and egg laying in Caenorhabditis elegans. Four distinct 5-HT receptors have been partially characterized, but little is known about their function in vivo. SER-7 exhibits most sequence identity to the mammalian 5-HT 7 receptors and couples to a stimulation of adenyl cyclase when expressed in COS-7 cells. However, many 5-HT 7 -specific agonists have low affinity for SER-7. 5-HT fails to stimulate pharyngeal pumping and the firing of the MC motorneurons in animals containing the putative ser-7(tm1325) and ser-7(tm1728) null alleles. In addition, although pumping on bacteria is upregulated in ser-7(tm1325) animals, pumping is more irregular. A similar failure to maintain ''fast pumping'' on bacteria also was observed in ser-1(ok345) and tph-1(mg280) animals that contain putative null alleles of a 5-HT 2 -like receptor and tryptophan hydroxylase, respectively, suggesting that serotonergic signaling, although not essential for the upregulation of pumping on bacteria, ''fine tunes'' the process. 5-HT also fails to stimulate egg laying in ser-7(tm1325), ser-1(ok345), and ser-7(tm1325) ser-1(ok345) animals, but only the ser-7 ser-1 double mutants exhibit an Egl phenotype. All of the SER-7 mutant phenotypes are rescued by the expression of full-length ser-7Tgfp translational fusions. ser-7Tgfp is expressed in several pharyngeal neurons, including the MC, M2, M3, M4, and M5, and in vulval muscle. Interestingly, 5-HT inhibits egg laying and pharyngeal pumping in ser-7 null mutants and the 5-HT inhibition of egg laying, but not pumping, is abolished in ser-7(tm1325);ser-4(ok512) double mutants. Taken together, these results suggest that SER-7 is essential for the 5-HT stimulation of both egg laying and pharyngeal pumping, but that other signaling pathways can probably fulfill similar roles in vivo.

Biogenic amines modulate key behaviors in both vertebrates and invertebrates. In Caenorhabditis elegans, tyramine (TA) and octopamine (OA) inhibit aversive responses to 100%, but not dilute (30%) octanol. TA and OA also abolish food-and serotonin-dependent increases in responses to dilute octanol in wild-type but not tyra-3(ok325) and f14d12.6(ok371) null animals, respectively, suggesting that TA and OA modulated responses to dilute octanol are mediated by separate, previously uncharacterized, G-protein-coupled receptors. TA and OA are high-affinity ligands for TYRA-3 and F14D12.6, respectively, based on their pharmacological characterization after heterologous expression. f14d12.6::gfp is expressed in the ASHs, the neurons responsible for sensitivity to dilute octanol, and the sra-6-dependent expression of F14D12.6 in the ASHs is sufficient to rescue OA sensitivity in f14d12.6(ok371) null animals. In contrast, tyra-3::gfp appears not to be expressed in the ASHs, but instead in other neurons, including the dopaminergic CEP/ADEs. However, although dopamine (DA) also inhibits 5-HT-dependent responses to dilute octanol, TA still inhibits in dop-2; dop-1; dop-3 animals that do not respond to DA and cat-2(tm346) and Pdat-1::ICE animals that lack significant dopaminergic signaling, suggesting that DA is not an intermediate in TA inhibition. Finally, responses to TA and OA selectively desensitize after preexposure to the amines. Our data suggest that although tyraminergic and octopaminergic signaling yield identical phenotypes in these olfactory assays, they act independently through distinct receptors to modulate the ASH-mediated locomotory circuit and that C. elegans is a useful model to study the aminergic modulation of sensory-mediated locomotory behaviors.

Serotonin (5-HT) regulates key processes in both vertebrates and invertebrates. Previously, four 5-HT receptors that contributed to the 5-HT modulation of egg laying were identified in Caenorhabditis elegans. Therefore, to assess potential receptor interactions, we generated animals containing combinations of null alleles for each receptor, especially animals expressing only individual 5-HT receptors. 5-HTstimulated egg laying and egg retention correlated well with different combinations of predicted excitatory and inhibitory serotonergic inputs. For example, 5-HT did not stimulate egg laying in ser-1, ser-7, or ser-7 ser-1 null animals, and ser-7 ser-1 animals retained more eggs than wild-type animals. In contrast, 5-HT-stimulated egg laying in ser-4;mod-1 animals was greater than in wild-type animals, and ser-4;mod-1 animals retained fewer eggs than wild-type animals. Surprisingly, ser-4;mod-1;ser-7 ser-1 animals retained the same number of eggs as wild-type animals and exhibited significant 5-HT-stimulated egg laying that was dependent on a previously uncharacterized receptor, SER-5. 5-HT-stimulated egg laying was absent in ser-5;ser-4;mod-1;ser-7 ser-1 animals, and these animals retained more eggs than either wild-type or ser-4;mod-1;ser-7 ser-1 animals. The 5-HT sensitivity of egg laying could be restored by ser-5 muscle expression. Together, these results highlight the dual excitatory/inhibitory serotonergic inputs that combine to modulate egg laying.

No abstract

Early development of the parasitic nematode, Ascaris suum, occurs inside a highly resistant eggshell, and the developing larva is bathed in perivitelline fluid. Twodimensional gel analysis of perivitelline fluid from infective larvae reveals seven major proteins; a cDNA encoding one of these, As-p18, has been cloned, sequenced, and protein expressed in Escherichia coli. The predicted amino acid sequence of As-p18 exhibits similarities to the intracellular lipid-binding protein (iLBP) family including retinoid-and fatty acid-binding proteins (FABP). As-p18 is unusual in that it possesses a hydrophobic leader that is not present in the mature protein, the developmental regulation of its expression, and in terms of its predicted structure.Recombinant As-p18 is a functional FABP with a high affinity for both a fluorescent fatty acid analog (11-(((5-(dimethylamino)-1-naphthalenyl )sulfonyl ) amino ) undecanoic acid) and oleic acid, but not retinol. Circular dichroism of rAs-p18 reveals a high ␤-sheet content (62%), which is consistent with secondary structure for the protein predicted from sequence algorithms, and the structure of iLBPs. Unusual features are apparent in a structural model of As-p18 generated from existing crystal structures of iLBPs.As-p18 is not found in unembryonated eggs, begins to be synthesized at about day 3 of development, reaches a maximal concentration with the formation of the firststage larva and remains abundant in the perivitelline fluid of the second-stage larva. Since As-p18 is not present in the post-infective third-stage larva or adult worm tissues, it appears to be exclusive to the egg. Surprisingly, however, Northern blot analysis yields mRNA for As-p18 not only in the early larval stages, but also the unembryonated egg, third-stage larvae, and ovaries of adult worms, even though the protein is not detectable from any of those sources. As-p18 may play a role in sequestering potentially toxic fatty acids and their peroxidation products, or it may be involved in the maintenance of the impermeable lipid layer of the eggshell.Parasitic nematodes cause medical and economic damage on a global scale, in part the result of the longevity and environmental robustness of their dispersal stages, their eggs. Arguably the most successful and widely distributed nematode parasite of humans is Ascaris lumbricoides, which infects over 1 billion people. A morphologically indistinguishable species, Ascaris suum, occurs in pigs and has become the best understood nematode in biochemical terms. The eggs contaminate soil, and the embryos develop to infective, second-stage larvae (L2s)