Acute effects of Pregabalin on the distribution and electrophysiological properties of the Cav2.1 calcium channels expressed in the HEK293t cell line

WEISSMANN CARINA; URBANO FRANCISCO J.; UCHITEL OSVALDO D.

Congreso; XXVI Congreso anual SAN.; 2011

We established a cell model to study the acute effects of Pregabalin (PGB), a drug widely used in epilepsy and neuropathic pain, on calcium channels at the cellular level. HEK293t cells were transfected with plasmids coding for all subunits of the Cav2.1 channel. We used aα1 fused to eGFP tag, whereas in parallel experiments α2δ subunits were visualized through a pHluorin tag to follow the distribution # 117 of the different subunits in time and at different experimental conditions. The expressed channel was functional as shown by increases in calcium currents of transfected cells measured by whole cell patch-clamp recordings, showing a maximum barium currents peak in the I-V curve at ~20 mV. The fluorescent signal showed a homogenous distribution; and acidification of the medium reduced the fluorescence when pHluorin- α2δ was expressed, which recovered after basification. Incubation of cells with 500 uM PGB for 30 minutes induced changes in localization of the subunits as measured by fluorescent time lapse microscopy, whereas no changes were visualized for cells expressing a control GFP protein. Together these results show strong evidence for an acute effect of PGB on calcium channels¥distribution and electrophysiological properties of the channel.