Article The Sensory Circuitry for Sexual Attraction in C. elegans Males (2007)

Department of BiologyUniversity of Utah257 South 1400 EastSalt Lake City, Utah 84112-0840SummaryBackground: Why do males and females behave differ-ently? Sexually dimorphic behaviors could arise fromsex-speciﬁc neurons or by the modiﬁcation of circuitspresent in both sexes. C. elegansmales exhibit differentbehaviors than hermaphrodites. Although there is a sin-gle class of sex-speciﬁc sensory neurons in the head ofmales, most of their neurons are part of a core nervoussystem also present in hermaphrodites. Are the behav-ioral differences due to sex-speciﬁc or core neurons?Results: We demonstrate that C. elegansmales chemo-tax to a source of hermaphrodite pheromones. This sex-ual-attraction behavior depends on a TRPV (transientreceptor potential vanilloid) channel encoded by theosm-9,ocr-1,andocr-2genes.OSM-9isrequiredinthreeclasses of sensoryneurons: the AWA and AWC olfactoryneurons and the male-speciﬁc CEM neurons. The ab-senceofOSM-9fromanyoftheseneuronsimpairsattrac-tion, suggesting that their ensemble output elicits sexualattraction. Likewise, the ablation of any of these classesafter sexualmaturationimpairs attractionbehavior. Ifab-lationsareperformedbeforesexualmaturation,attractionis unimpaired, demonstrating that these neurons com-pensateforoneanother.Thus,maleslackingsex-speciﬁcneurons are still attracted to pheromones, suggestingthat core neurons are sexualized. Similarly, transgendernematodes—animals that appear morphologically to behermaphrodites but have a masculinized core nervoussystem—are attracted to hermaphrodite pheromones.Conclusions: Both sexually dimorphic and core sen-sory neurons are normally required in the adult for sex-ual attraction, but they can replace each other duringsexual maturation if necessary to generate robust male-speciﬁc sexual attraction behavior.IntroductionIn the hermaphroditic nematode C. elegans, it is incum-bent on the male to ﬁnd a mate. There is considerableevidence for male-speciﬁc sexual attraction in nema-todes (reviewed in [1–3]). C. elegans males remain on afood source if a hermaphrodite is present [4] and re-spond to cues released speciﬁcally by hermaphrodites[5]–clear evidence for male-speciﬁc chemosensorybehavior. What are the differences between the maleandhermaphrodite nervous systemsthat could accountfor sexually dimorphic behaviors? At ﬁrst glance, thenervous systems of hermaphrodites and males are rad-ically different. Out of 383 total neurons in males, 89 aresex-speciﬁc.However,mostofthesex-speciﬁcneuronsin themale innervate thespecialized copulatory appara-tus of the male, the male tail. Thus, much of the male-speciﬁc circuitry is likely to be involved in copulationrather than chemotaxis to hermaphrodites [6]. Theremaining 294 neurons in the male comprise a corenervous system also found in hermaphrodites [7]. All ofthesensoryneuronsinthehead,responsibleforchemo-taxis of hermaphrodites to simple compounds, are alsofound in males. Consistent with this anatomical similar-ity, the chemosensory responses of the two sexes tosimple compounds are to a ﬁrst approximation thesame [8]. In addition, males have four sex-speciﬁcchemosensory neurons in the head, the CEM neurons.These neurons undergo programmed cell death duringhermaphrodite development, surviving only in males[7]. The male-speciﬁc CEMs have long been hypothe-sized to mediate chemosensory mate ﬁnding behaviorin C. elegans [9]. Thus, male-speciﬁc sexual attractioncould be due solely to the sex-speciﬁc CEM neurons,to subtle differences in the core circuitry, or a combina-tion of the two.ResultsMale Sexual-Attraction BehaviorC.elegansmalestax to a source of hermaphrodite pher-omones and tend to remain there (Figures 1 and 2A). Tocharacterize this response in C. elegans, we adapteda two-spot assay similar to those developed for studiesof chemotaxis [10–12]. This assay demonstrates that C.elegans males tax to the peak of a pheromone gradientand remain at the source (Figures 1 and 2A). To distin-guish the chemosensory behaviors elicited by phero-mones from behaviors requiring tactile input, we usedmedia conditioned by hermaphrodites (see the Supple-mental Experimental Procedures and Figure S2 in theSupplemental Data available online). To generate condi-tionedmedia,wegrewC.eleganshermaphroditesinliq-uid culture, removed the worms and bacteria food, andﬁlteredtheremainingliquid(SupplementalExperimentalProcedures). Drops of conditioned and unconditionedmedia were placed on an agar test plate and allowedto soak in, and a thin coat of bacterial food was spreadover the entire surface. Males on a test plate locate thesource of hermaphrodite-conditioned media from a dis-tanceandtendtostaythere(Figures1,2A,and2B).Malesrespond to pheromones at a distance of up to 30 mm;analysis of single males (Figures 2C and 2D, Figure S1)shows that once an individual starts, he continuouslymoves toward the source until he is within 10 mm of thesourcecenter(Figure2D,FigureS1).Attractionisspeciﬁc