Abstract
Cyclic adenosine monophosphate (cAMP) has been implicated in the execution of diverse rhythmic behaviors, but how cAMP functions in neurons to generate behavioral outputs remains unclear. During the defecation motor program in C. elegans, a peptide released from the pacemaker (the intestine) rhythmically excites the GABAergic neurons that control enteric muscle contractions by activating a G protein-coupled receptor (GPCR) signaling pathway that is dependent on cAMP. Here, we show that the C. elegans PKA catalytic subunit, KIN-1, is the sole cAMP target in this pathway and that PKA is essential for enteric muscle contractions. Genetic analysis using cell-specific expression of dominant negative or constitutively active PKA transgenes reveals that knockdown of PKA activity in the GABAergic neurons blocks enteric muscle contractions, whereas constitutive PKA activation restores enteric muscle contractions to mutants defective in the peptidergic signaling pathway. Using real-time, in vivo calcium imaging, we find that PKA activity in the GABAergic neurons is essential for the generation of synaptic calcium transients that drive GABA release. In addition, constitutively active PKA increases the duration of calcium transients and causes ectopic calcium transients that can trigger out-of-phase enteric muscle contractions. Finally, we show that the voltage-gated calcium channels UNC-2 and EGL-19, but not CCA-1 function downstream of PKA to promote enteric muscle contractions and rhythmic calcium influx in the GABAergic neurons. Thus, our results suggest that PKA activates neurons during a rhythmic behavior by promoting presynaptic calcium influx through specific voltage-gated calcium channels.
Highlights
Cyclic adenosine monophosphate is a potent second messenger that plays an important role in cellular responses to extracellular signals to regulate a wide array of biological processes
CAMP is synthesized by adenylyl cyclases (ACs), which are activated by G protein-coupled receptors (GPCRs) that are coupled to the heterotrimeric G protein a subunit, Gas [9]
We identify protein kinase (PKA) as the major downstream target of Cyclic adenosine monophosphate (cAMP) in the NLP-40-AEX-2/GPCR peptidergic signaling pathway that functions in the GABAergic neurons to regulate the Exp step during the defecation motor program in C. elegans
Summary
Cyclic adenosine monophosphate (cAMP) is a potent second messenger that plays an important role in cellular responses to extracellular signals to regulate a wide array of biological processes. CAMP signaling is critical for the execution of rhythmic physiological processes such as heart beating and circadian rhythm in a variety of organisms [5,6,7,8]. Work in a variety of cell types has shown that cAMP has three major molecular targets: cyclic nucleotide-gated (CNG) channels, exchange proteins directly activated by cAMP (Epac) and cAMP-dependent protein kinase (PKA) (Figure 1A and [9]). PKA is a conserved serine/threonine kinase that has been implicated in a wide array of biological processes, including cell growth, neural function, cell differentiation and metabolism [12]. PKA activity has been implicated in the execution of rhythmic behaviors, such as sleep and circadian locomotor activity in the fly [14,15]
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