Intestinal signaling to GABAergic neurons regulates a rhythmic behavior in Caenorhabditis elegans

Abstract

The Caenorhabditis elegans defecation motor program (DMP) is a highly coordinated rhythmic behavior that requires two GABAergic neurons that synapse onto the enteric muscles. One class of DMP mutants, called anterior body wall muscle contraction and expulsion defective (aex) mutants, exhibits similar defects to those caused by the loss of these two neurons. Here, we demonstrate that aex-2 encodes a G-protein-coupled receptor (GPCR) and aex-4 encodes an exocytic SNAP25 homologue. We found that aex-2 functions in the nervous system and activates a Gs␣ signaling pathway to regulate defecation. aex-4, on the other hand, functions in the intestinal epithelial cells. Furthermore, we show that aex-5, which encodes a pro-protein convertase, functions in the intestine to regulate the DMP and that its secretion from the intestine is impaired in aex-4 mutants. Activation of the Gs␣ GPCR pathway in GABAergic neurons can suppress the defecation defect of the intestinal mutants aex-4 and aex-5. Lastly, we demonstrate that activation of GABAergic neurons using the light-gated cation channel channelrhodopsin-2 is sufficient to suppress the behavioral defects of aex-2, aex-4, and aex-5. These results genetically place intestinal genes aex-4 and aex-5 upstream of GABAergic GPCR signaling. We propose a model whereby the intestinal genes aex-4 and aex-5 control the DMP by regulating the secretion of a signal, which activates the neuronal receptor aex-2. T he Caenorhabditis elegans defecation motor program (DMP)is a highly coordinated series of three muscle contractions that are executed every 45 sec [ Fig. 1A and supporting information (SI) Movie S1]. The cycle is initiated by a posterior body wall muscle contraction (pBoc), followed 2-3 sec later by an anterior body wall muscle contraction (aBoc). About 1 sec after the aBoc, enteric muscles contract, thus causing the expulsion (Exp) of intestinal contents. The process repeats itself Ϸ45 sec later with little variability in the timing of contractions (1). A genetic screen for mutants that displayed defects in the DMP isolated mutants defective in each of the three muscle contractions, known as pbo, abo, and exp (1). The screen also recovered mutants defective in the last two muscle contractions (aBoc and Exp [aex]) and mutants defective in the cycle periodicity (i.e., longer or shorter than normal DMP cycling times) (1). Molecular studies of these mutants have suggested that the behavior is orchestrated through the communication between the intestine, GABAergic neurons, and muscle.The periodicity of the DMP is regulated by the C. elegans intestine, a single-cell layer tube of polarized epithelial cells joined by gap junctions (2, 3). Intestinal Ca 2ϩ oscillations with Ϸ45-sec periodicity appear to play a central role in this timing. They consist of a posterior-to-anterior Ca 2ϩ wave whose levels peak in the posterior and anterior intestinal cells just before the pBoc and aBoc contractions, respectively (3-5). Mutations in genes involved in the maintenance of Ca 2ϩ oscillations or i...

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“…There are numerous developmental and behavioural phenomena that rely on communication between the nervous system and intestinal tissue in C. elegans. For example, neuron-intestine communication via insulin-like peptides regulates the dauer diapause decision during development 25 , and intestinal signalling to GABAergic neurons regulates a highly coordinated defecation motor programme 26 . Furthermore, we previously reported a cell-nonautonomous effect for mutant TDP-43 proteins in activating the ER unfolded protein response.…”

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“…EG3680 oxIs206[aex-3 p ANF-GFP] was kindly provided by Erik Jorgensen (Speese et al 2007). NM3576 jsIs1072 (vha-6 p -AEX-5-VENUS] was previously described (Mahoney et al 2008). KG2361 nuIs183 [unc-129 -NLP-21-Venus] was previously described (Sieburth et al 2007).…”

“…aex-3 p -ANF-GFP imaging experiments of the dorsal nerve cord and coelomocytes were carried out using an Olympus BX51, equipped with a digital camera Micropublisher 3.3 RTV ( JH Technologies, San Francisco). Images of coelomocytes from animals expressing atrial natriuretic factor (ANF)-GFP were taken and analyzed as previously described (Mahoney et al 2008). Images of rab3 p -HID-1-GFP and aex-3 p -ANF-GFP in the dorsal nerve cord were captured using a 360 oil objective.…”

“…Images of aex-3 p -ANF-GFP in coelomocytes were captured using a 340 objective. Quantification of AEX-5-VENUS was performed as described previously (Mahoney et al 2008).…”

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