Dissociation of the G protein ßγ from the Gq-PLCß complex partially attenuates PIP2 hydrolysis.

Phospholipase C ß (PLCß), which is activated by the Gq family of heterotrimeric G proteins, hydrolyzes the inner membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2), generating diacylglycerol and inositol 1,4,5-triphosphate (IP3). Because Gq and PLCß regulate many crucial cellular processes and have been identified as major disease drivers, activation and termination of PLCß signaling by the Gαq subunit have been extensively studied. Gq-coupled receptor activation induces intense and transient PIP2 hydrolysis, which subsequently recovers to a low-intensity steady-state equilibrium. However, the molecular underpinnings of this equilibrium remain unclear. Here, we explored the influence of signaling crosstalk between Gq and Gi/o pathways on PIP2 metabolism in living cells using single-cell and optogenetic approaches to spatially and temporally constrain signaling. Our data suggest that the Gßγ complex is a component of the highly efficient lipase GαqGTP-PLCß-Gßγ. We found that over time, Gßγ dissociates from this lipase complex, leaving the less-efficient GαqGTP-PLCß lipase complex and allowing the significant partial recovery of PIP2 levels. Our findings also indicate that the subtype of the Gγ subunit in Gßγ fine-tunes the lipase activity of Gq-PLCß, in which cells expressing Gγ with higher plasma membrane interaction show lower PIP2 recovery. Given that Gγ shows cell- and tissue-specific subtype expression, our findings suggest the existence of tissue-specific distinct Gq-PLCß signaling paradigms. Furthermore, these results also outline a molecular process that likely safeguards cells from excessive Gq signaling.

Confocal imaging of cytosolic Ca2+ and fuzzy clustering reveal the circuit topology details underlying synchronization in hippocampal neurons.

Neuronal synchronization contributes to various cognitive functions and disruption in synchronicity may lead to various diseased conditions. However, measurement of synchronicity at a higher spatial resolution remains challenging. Specifically, investigation on understanding the role of network topology in tuning the network activity and synchronicity remains sparse. In this context, we propose imaging of intracellular Ca2+ in primary cultures of hippocampal neurons using Fluo-4 as the fluorescent indicator using the confocal microscope. In order to identify the synchronous response from a set of heterogeneous Ca2+ spiking, we present fuzzy clustering of the oscillatory responses. Further, the synchronicity was measured through evaluation of the correlation between Ca2+ spiking trends. Confocal imaging and analysis show that neuronal connectivity and topology play an essential role in tuning the synchronicity of the neuronal network.