Functional interaction of auxiliary subunits and synaptic proteins with Ca_V1.3 May impart hair cell Ca²⁺ current properties

We assessed the functional determinants of the properties of L-type Ca²⁺ currents in hair cells by co-expressing the pore-forming Ca_v1.3α₁ subunit with the auxiliary subunits β_1A and/or α_2δ. Because Ca²⁺ channels in hair cells are poised to interact with synaptic proteins, we also co-expressed the Ca_v1.3α₁ subunit with syntaxin, vesicle-associated membrane protein (VAMP), and synaptosome associated protein of 25 kDa (SNAP25). Expression of the Ca_v1.3α₁ subunit in human embryonic kidney cells (HEK 293) produced a dihydropyridine (DHP)-sensitive Ca²⁺ current (peak current density - 2.0 ± 0.2 pA/pF; n = 11). Co-expression with β_1A and α_2δ, subunits enhanced the magnitude of the current (peak current density: Ca_v1.3α₁ + β_1A = -4.3 ± 0.8 pA/pF, n = 10; Ca_v1.3α₁ + β_1A + α_2δ = -4.1 ± 0.6 pA/pF, n = 9) and produced a leftward shift of approximately 9 mV in the voltage-dependent activation of the currents. Furthermore, co-expression of Ca_v1.3α₁ with syntaxin/ VAMP/SNAP resulted in at least a twofold increase in the peak current density (-4.7 ± 0.2 pA/pF; n = 11) and reduced the extent of inactivation of the Ca²⁺ currents. Botulinum toxin, an inhibitor of syntaxin, accelerated the inactivation profile of Ca²⁺ currents in hair cells. Immunocytochemical data also indicated that the Ca²⁺ channels and syntaxin are co-localized in hair cells, suggesting there is functional interaction of the Ca_v1.3α₁ with auxiliary subunits and synaptic proteins, that may contribute to the distinct properties of the DHP-sensitive channels in hair cells.