Presynaptic Gα_o(GOA-1) signals to depress command neuron excitability and allow stretch-dependent modulation of egg laying inCaenorhabditis elegans

Abstract

27Caenorhabditis elegans egg laying is a two-state behavior modulated by sensory input. 28Feedback of egg accumulation in the uterus drives activity of the serotonergic HSN command 29 neurons to promote the active state, but how aversive sensory stimuli signal to inhibit egg laying 30 is not well understood. We find the Pertussis Toxin-sensitive G protein, Go, signals in HSN to 31 inhibit circuit activity and prolong the inactive behavior state. Go signaling hyperpolarizes HSN, 32reducing Ca 2+ activity and input into the postsynaptic vulval muscles. Loss of inhibitory Go 33 signaling uncouples presynaptic HSN activity from a postsynaptic, stretch-dependent homeostat, 34 causing precocious entry into the egg-laying active state. NLP-7 neuropeptides signal to reduce 35 egg laying both by inhibiting HSN and by activating Go in cells other than HSN. Thus, Go 36integrates diverse signals to maintain a bi-stable state of electrical excitability that dynamically 37 controls circuit activity and behavior output in response to a changing environment. 38 . 39 40 41 42 43 44 45 46 We have recently identified a stretch-dependent homeostat that scales egg-laying circuit 95 activity in response to feedback of egg accumulation. Juvenile and young adult animals lacking 96 eggs in the uterus have low circuit activity, and optogenetic stimulation of the HSNs is unable to 97 stimulate vulval muscle activity in these animals (Ravi et al., 2018a). Chemical or genetic 98 sterilization leads to a reduction in both HSN and vulval muscle Ca 2+ activity, locking animals in 99 the inactive state (Collins et al., 2016; Ravi et al., 2018a). Acute chemogenetic silencing of vulval 100 muscle electrical activity similarly blocks egg laying and presynaptic HSN Ca 2+ activity. Reversal 101 of this muscle silencing drives a homeostatic rebound in HSN 'burst' firing Ca 2+ activity where 102 'bursts' of HSN Ca 2+ transients promote ongoing circuit activity that drives release of the excess 103 accumulated eggs (Ravi et al., 2018a). Feedback of successful egg release also signals to inhibit 104 HSN activity. Four uv1 neuroendocrine cells which line the vulval canal are mechanically 105 activated by the passage of eggs. The uv1 cells are peptidergic and tyraminergic, and inhibition 106 of egg laying by tyramine requires the LGC-55 tyramine-gated Clchannel which is expressed 107 on the HSNs (Collins et al., 2016). uv1 also expresses the FLP-11 and NLP-7 neuropeptides 108 that signal to inhibit HSN activity and egg laying through receptors that remain unidentified 109 (Banerjee et al., 2017). Full NLP-7 inhibition of egg laying requires the EGL-47 receptor and the 110 G protein, Go, both of which are expressed in HSN (Moresco and Koelle, 2004; Banerjee et al., 111 2017). HSN Ca 2+ activity and egg laying are also inhibited by aversive signals from the external 112 environment. Elevated environmental CO2 activates BAG and other sensory neurons (Hallem et 113 al., 2011;Fenk and de Bono, 2015). BAG releases FLP-17, which binds to EGL-6 recepto...

Serotonin signals through postsynaptic Gαq, Trio RhoGEF, and diacylglycerol to promoteC. elegansegg-laying circuit activity and behavior

“…In C. elegans , 2-AG activates the NPR-19 endocannabinoid receptor ortholog that couples to G α o to modulate serotonin transmission, pharyngeal, feeding, and locomotory behaviors (Oakes et al, 2019; Oakes et al, 2017; Pastuhov et al, 2016). We have recently shown that feedback of egg accumulation alters vulval muscle Ca 2+ activity, which subsequently signals to regulate bursting in the HSNs (Ravi et al, 2018b; Ravi et al, 2020), These results support a model where stretch-dependent feedback of egg accumulation stimulates postsynaptic vulval muscle Ca 2+ signaling. This Ca 2+ would then activate PLCs to generate DAG and 2-AG which signal to modulate HSN activity, serotonin release, and egg laying.…”

“…The copyright holder for this preprint (which this version posted May 10, 2021. ; https://doi.org/10.1101/2021.05.08.443256 doi: bioRxiv preprint result of the shortness of the 10-minute Ca 2+ recording period that followed PMA treatment. Egglaying active states typically occur every ~20 minutes in wild-type animals (Waggoner et al, 1998) and every 12-15 minutes in hyperactive egg-laying behavior mutants that resemble the effects of PMA treatment (Ravi et al, 2020;Waggoner et al, 2000b). The M9 buffer assays allow egg-laying events to accumulate over one hour without having to capture specific events.…”

“…In C. elegans, 2-AG activates the NPR-19 endocannabinoid receptor ortholog that couples to Gao to modulate serotonin transmission, pharyngeal, feeding, and locomotory behaviors (Oakes et al, 2019;Oakes et al, 2017;Pastuhov et al, 2016). We have recently shown that feedback of egg accumulation alters vulval muscle Ca 2+ activity, which subsequently signals to regulate bursting in the HSNs (Ravi et al, 2018b;Ravi et al, 2020),…”

The neural G protein Gαo tagged with GFP at an internal loop is functional in Caenorhabditis elegans

“…Caenorhabditis elegans has a Gα o ortholog named GOA-1 that is >80% identical to mammalian Gα o and that is expressed in most or all neurons. GOA-1 has been shown by genetic analysis to inhibit neurotransmitter release and/or neural activity ( Mendel et al 1995 ; Ségalat et al 1995 ; Nurrish et al 1999 , Ravi et al 2020 ), but the molecular mechanisms by which Gα o signals to have these effects remain to be fully defined. While activated Gα o releases Gβγ subunits to regulate specific potassium and calcium channels ( Lüscher and Slesinger 2010 ; Proft and Weiss 2015 ), genetic studies in C. elegans suggest that signaling through Gβγ is not likely the sole mechanism by which Gα o has its physiological effects ( Koelle 2018 ).…”

The neural G protein Gαotagged with GFP at an internal loop is functional inC. elegans

“…C. elegans has a Gα o ortholog named GOA-1 that is >80% identical to mammalian Gα o and that is expressed in most or all neurons. GOA-1 has been shown by genetic analysis to inhibit neurotransmitter release and/or neural activity (Mendel et al, 1995; Ségalat et al, 1995; Nurrish et al 1999, Ravi et al, 2020), but the molecular mechanisms by which Gα o signals to have these effects remain to be fully defined. While activated Gα o releases Gβγ subunits to regulate specific potassium and calcium channels (Lüscher and Slesinger, 2010; Proft and Weiss, 2015), genetic studies in C. elegans suggest that signaling through Gβγ is not likely the sole mechanism by which Gα o has its physiological effects (Koelle, 2018).…”