Proprioceptors are low-threshold mechanoreceptors involved in perceiving body position and strain bearing. However, the physiological response of proprioceptors to fatigue- and muscle-acidosis-related disturbances remains unknown. Here, we employed whole-cell patch-clamp recordings to probe the effect of mild acidosis on the mechanosensitivity of the proprioceptive neurons of dorsal root ganglia (DRG) in mice. We cultured neurite-bearing parvalbumin-positive (Pv+) DRG neurons on a laminin-coated elastic substrate and examined mechanically activated currents induced through substrate deformation-driven neurite stretch (SDNS). The SDNS-induced inward currents (I_SDNS) were indentation depth-dependent and significantly inhibited by mild acidification (pH 7.2~6.8). The acid-inhibiting effect occurred in neurons with an I_SDNS sensitive to APETx2 (an ASIC3-selective antagonist) inhibition, but not in those with an I_SNDS resistant to APETx2. Detailed subgroup analyses revealed I_SDNS was expressed in 59% (25/42) of Parvalbumin-positive (Pv+) DRG neurons, 90% of which were inhibited by APETx2. In contrast, an acid (pH 6.8)-induced current (I_Acid) was expressed in 76% (32/42) of Pv+ DRG neurons, 59% (21/32) of which were inhibited by APETx2. Together, ASIC3-containing channels are highly heterogenous and differentially contribute to the I_SNDS and I_Acid among Pv+ proprioceptors. In conclusion, our findings highlight the importance of ASIC3-containing ion channels in the physiological response of proprioceptors to acidic environments.

Ca2+ influx through voltage-gated Ca2+ channels (CaVs) at the plasma membrane is the major pathway responsible for the elevation of the intracellular Ca2+ concentration ([Ca2+]i), which activates various physiological activities. Calmodulin (CaM) is known to be involved in the Ca2+-dependent inactivation (CDI) of several types of CaVs; however, little is known about how CaM modulates CaV2.2. Here, we expressed CaV2.2 with CaM or CaM mutants with a Ca2+-binding deficiency in HEK293T cells and measured the currents to characterize the CDI. The results showed that CaV2.2 displayed a fast inactivation with Ca2+ but not Ba2+ as the charge carrier; when CaV2.2 was co-expressed with CaM mutants with a Ca2+-binding deficiency, the level of inactivation decreased. Using glutathione S-transferase-tagged CaM or CaM mutants as the bait, we found that CaM could interact with the intracellular C-terminal fragment of CaV2.2 in the presence or absence of Ca2+. However, CaM and its mutants could not interact with this fragment when mutations were generated in the conserved amino acid residues of the CaM-binding site. CaV2.2 with mutations in the CaM-binding site showed a greatly reduced current that could be rescued by CaM12 (Ca2+-binding deficiency at the N-lobe) overexpression; in addition, CaM12 enhanced the total expression level of CaV2.2, but the ratio of CaV2.2 present in the membrane to the total fraction remained unchanged. Together, our data suggest that CaM, with different Ca2+-binding abilities, modulates not only the inactivation of CaV2.2 but also its expression to regulate Ca2+-related physiological activities.