Store-operated calcium entry in vagal sensory nerves is independent of Orai channels

摘要

Vagal sensory nerves innervate the majority of visceral organs (e.g. heart, lungs, GI tract, etc) and their activation is critical for defensive and regulatory reflexes. Intracellular Ca²⁺ is a key regulator of neuronal excitability and is largely controlled by the Ca²⁺ stores of the endoplasmic reticulum. In other cell types store-operated channels (SOC) have been shown to contribute to the homeostatic control of intracellular Ca²⁺. Here, using Ca²⁺ imaging, we have shown that ER depletion in vagal sensory neurons (using thapsigargin or caffeine) in the absence of extracellular Ca²⁺ evoked Ca²⁺ influx upon reintroduction of Ca²⁺ into the extracellular buffer. This store-operated Ca²⁺ entry (SOCE) was observed in approximately 25–40% of vagal neurons, equally distributed among nociceptive and non-nociceptive sensory subtypes. SOCE was blocked by Gd³⁺ but not by the Orai channel blocker . We found Orai channel mRNA in extracts from whole vagal ganglia, but when using single cell RT-PCR analysis we found only 3 out of 34 neurons expressed Orai channel mRNA, indicating that Orai channel expression in the vagal ganglia was likely derived from non-neuronal cell types. Confocal microscopy of vagal neurons in 3 day cultures demonstrated rich ER tracker fluorescence throughout axonal and neurite structures and ER store depletion (thapsigargin) evoked Ca²⁺ transients from these structures. However, no SOCE could be detected in the axonaleurite structures of vagal neurons. We conclude that SOCE occurs in vagal sensory neuronal cell bodies through non-Orai mechanisms but is absent at nerve terminals.