LAUSR

Calcium (Ca2+) plays a pivotal role in modulating neuronal-mediated responses to multimodal sensory stimuli. Recent studies in Drosophila reveal class III (CIII) multidendritic (md) sensory neurons function as multimodal sensors regulating distinct behavioral responses to innocuous mechanical and nociceptive thermal stimuli. Functional analyses indicate that CIII-mediated multimodal behavioral output is dependent upon activation levels with stimulus-evoked Ca2+ displaying relatively low vs. high intracellular levels in response to gentle touch vs. noxious cold, respectively. However, the mechanistic bases underlying modality-specific differential Ca2+ responses in CIII neurons remain incompletely understood. We hypothesized that noxious cold-evoked high intracellular Ca2+ responses in CIII neurons may rely upon Ca2+-induced Ca2+ release (CICR) mechanisms involving transient receptor potential (TRP) channels and/or metabotropic G-protein coupled receptor (GPCR) activation to promote cold nociceptive behaviors. GABAB receptor mutants and CIII-specific knockdown resulted in impaired noxious cold-evoked behaviors. Gαq and Phospholipase C signaling are likewise required for noxious cold sensing. Additionally, ER localized Ca2+ channels including the Ryanodine receptor (RyR) and Inositol trisphosphate receptor (IP3R) are required for cold nociceptive behaviors. GPCR mediated signaling, through GABAB-R2 and IP3R, is not required in CIII neurons for innocuous touch evoked behaviors. However, CICR via RyR is required in CIII neurons for innocuous touch-evoked behaviors. Disruptions in GABAB -R2, IP3R and RyR in CIII neurons leads to significantly lower levels of cold-evoked Ca2+ responses indicating GPCR and CICR signaling mechanisms function in regulating Ca2+ release. CIII neurons exhibit bipartite cold-evoked firing patterns, where CIII neurons burst during rapid temperature change and tonically fire during steady state cold temperatures. GABAB-R2 knockdown in CIII neurons resulted in disorganized firing patterns during cold exposure. Upon ryanodine pharmacological application, CIII neurons exhibit increased bursting activity and with CIII specific RyR knockdown, there is an increase in cold-evoked tonic firing and decrease in bursting. Lastly, our previous studies implicated the TRPP channel Pkd2 in cold nociception, and here, we show that Pkd2 and IP3R genetically interact in regulating cold-evoked behavior. Collectively, these analyses support novel, modality-specific roles for metabotropic GABAergic signaling and CICR mechanisms in regulating intracellular Ca2+ levels and cold-evoked behavioral output from multimodal CIII neurons.