Figure 7.

NLP-7 neuropeptides signal through EGL-47 and Gα_o to inhibit egg-laying outside of the HSNs. (A) NLP-7 signals through Gα_o to inhibit egg-laying behavior. Scatter plots show average number of eggs retained by wild-type (gray circles), goa-1(sa734) mutants (blue open circles), NLP-7 over-expressing (OE) transgenics in the wild-type (pink triangles or orange squares), and in NLP-7 over-expressing transgenics in the goa-1(n1134) (pink open triangles) and goa-1(sa734) null mutant background (orange open squares). Data in orange squares are from animals that also carry the vsIs183 transgene used for HSN Ca²⁺ imaging. Mean totals below ∼10 eggs indicate hyperactive egg laying while totals above ∼20 eggs indicate egg-laying defective behavior defects. Error bars indicate 95% confidence intervals for the mean. Asterisks indicate P < 0.0001 (one-way ANOVA with Bonferroni correction for multiple comparisons). N ≥ 30 animals for each strain. (B) Measure of early stage eggs laid by wild-type (black), goa-1(sa734) null mutants (blue), and goa-1(sa734) null mutants over-expressing NLP-7 neuropeptides (orange). Both goa-1(sa734) mutant strains also carry the vsIs183 transgene used for HSN Ca²⁺ imaging. (C) NLP-7 over-expression silences HSN Ca²⁺ activity. Representative GCaMP5/mCherry ratio traces showing HSN Ca²⁺ activity in wild-type (black), goa-1(sa734) null mutants, and NLP-7 overexpressing transgenic animals in the wild-type (pink) and goa-1(sa734) null mutant backgrounds (orange). Arrowheads indicate egg-laying events. (D) Scatter plots show HSN Ca²⁺ peaks per minute measurements in wild-type (black), goa-1(sa734) null mutants, or egl-47(ok677) null mutant animals in the absence or presence of NLP-7 over-expression (OE). Error bars indicate 95% confidence intervals for the mean for N ≥ 10 animals; n.s. indicates P > 0.05 while asterisk indicates P ≤ 0.0001 (one-way ANOVA with Bonferroni correction for multiple comparisons). (E) NLP-7 inhibition of egg laying is not exclusive to HSN silencing. Scatter plots show the average number of eggs retained by wild-type animals, transgenic animals expressing Pertussis Toxin in HSN, and egl-47(ok677) mutant animals in the absence or presence of NLP-7 over-expression (OE) or the absence of HSNs. Error bars indicate 95% confidence intervals for the mean. Asterisks indicate P < 0.0001 (one-way ANOVA with Bonferroni correction for multiple comparisons); N ≥ 30 animals for each strain. (F) Measure of early stage eggs laid by wild-type or transgenic animals expressing Tetanus Toxin the VC neurons (black outlined boxes) in animals expressing Pertussis Toxin in the HSNs (green) or in goa-1(sa734) animals missing Gα_o (blue). Error bars indicate ± 95% confidence intervals for the mean proportion. Asterisk indicates highlighted significant differences (P ≤ 0.0001) while n.s. indicates P > 0.05 (Fisher Exact Test with Bonferroni correction for multiple comparisons).