Functional expression of L-, N-, P/Q-, and R-type calcium channels in the human NT2-N cell line.

摘要

The biophysical and pharmacological properties of voltage-gated calcium channel currents in the human teratocarcinoma cell line NT2-N were studied using the whole cell patch-clamp technique. When held at -80 mV, barium currents (I(Ba)s) were evoked by voltage commands to above -35 mV that peaked at +5 mV. When holding potentials were reduced to -20 mV or 5 mM barium was substituted for 5 mM calcium, there was a reduction in peak currents and a right shift in the current-voltage curve. A steady-state inactivation curve for I(Ba) was fit with a Boltzmann curve (V(1/2) = -43.3 mV; slope = -17.7 mV). Maximal current amplitude increased from 1-wk (232 pA) to 9-wk (1025 pA) postdifferentiation. Whole cell I(Ba)s were partially blocked by specific channel blockers to a similar extent in 1- to 3-wk and 7- to 9-wk postdifferentiation NT2-N cells: 10 microM nifedipine (19 vs. 25%), 10 microM conotoxin GVIA (27 vs. 25%), 10 microM conotoxin MVIIC (15 vs. 16%), and 1.75 microM SNX-482 (31 vs. 33%). Currents were completely blocked by 300 microM cadmium. In the presence of nifedipine, GVIA, and MVIIC, approximately 35% of current remained, which was reduced further by SNX-482 (7-14% of current remained), consistent with functional expression of L-, N-, and P/Q-calcium channel types and one or more R-type channel. The presence of multiple calcium currents in this human neuronal-type cell line provides a potentially useful model for study of the regulation, expression and cellular function of human derived calcium channel currents; in particular the R-type current(s).