PGB acutely affects large P/Q mediated currents and P/Q channel´s cellular distribution [Mo111]

WEISSMANN C; URBANO FJ; GONZALEZ INCHAUSPE C; DI GUILMI MN; UCHITEL OD

Cancun

Congreso; F.A.L.A.N. Congress National Congress of Physiological Sciences and Neuroscience and Neurobiology of Mexico; 2012

F.A.L.A.N. Congress National Congress of Physiological Sciences and Neuroscience and Neurobiology of Mexico

We set out to investigate the mechanism of action of pregabalin (PGB) -an anticonvulsant and analgesic drug used in the treatment of epilepsy and neuropathic pain- on P/Q-calcium channels.We previously reported that slices from calyx of Held of WT-mice incubated acutely with PGB (500 microM) show slightly reduced P/Q-calcium (CaV2.1) currents compared to untreated ones (Di Guilmi et al, 2010). In this work, replacing calcium by barium, we showed greater differences between PGB-treated and untreated slices (17±1.2% reduction), without any changes in voltage activating properties. We also analyzed HEK293t cells transfected with subunits of CaV2.1 channels, and detected by an eGFP-tagged alpha1 subunit. Fluorescent cells yielded robust barium-mediated currents that decreased (56±12%) after PGB (500 microM)treatment.Thus, at a real synapse as well as in an heterologous system, PGB applied acutely determined the reduction of large CaV2.1-mediated barium currents. To test whether the effect of PGB was dependent on the magnitude of CaV2.1-mediated currents, we analyzed the effect of PGB in transgenic R192Q KI mice. These mice express a familial hemiplegic migraine type-1 mutation andshow greater current amplitude during long lasting action potentials (APs) stimulation compared to WT animals (Inchauspe et al, 2010). KI slices incubated with PGB decreased the currents to WT-levels when stimulated with long duration APs (larger currents). This effect was absent with short APs, Using time-lapse fluorescence microscopy of HEK293t transfected cells. We were able to follow the distribution of the channels after acute incubation of PGB showing decreased (~30%) membrane fluorescence and suggesting the internalization of through acute PGB-mediated mechanism.These results not only provide the mechanism of action of PGB, but also highlight the importance of the drug as a potential candidate to use in other calcium channelopathies to drive currents to steady state levels.