Serum Factor Alters T-Type Cav3.2 Gating Kinetics and Current Density

Abstract

T-type Calcium channels play a critical role in regulating neuronal excitability and modulating sensory transmission. The Cav3.2 channel isoform is highly expressed in peripheral nociceptors as well as in the pain-processing regions of the dorsal horn of the spinal cord. Augmenting Cav3.2 currents has been shown to induce hyperexcitability in nociceptive neurons in vitro and hyperalgesia in vivo. These studies strongly suggest that potentiation of the Cav3.2 channel results in abnormal nociceptive transmission, which could contribute to a variety of clinical pain syndromes. Therefore, it is important to identify endogenously produced molecular species that modulate Cav3.2 currents. Using the patch-clamp technique and stably transfected human embryonic kidney cells (HEK-293) expressing the Cav3.2 channel, we have begun to characterize a factor found in fetal bovine serum (FBS) that profoundly affects Cav3.2 channel gating kinetics. Specifically, when compared to baseline recombinant currents, 1% serum produces maximal increases in current magnitude (350%; p<.001), conductance (150%, p<.001), rate of macroscopic inactivation (47.1%; p<.001) and deactivation (74.3%; p<.001). Furthermore, 1% serum induces a hyperpolarizing shift in voltage-dependence of activation (V50) (−4.77mV; p <.001) with minimal effect on voltage-dependence of inactivation. In contrast, we found that recombinant CaV3.1 currents were completely insensitive. Similar to recombinant Cav3.2 currents, T-currents from rat dorsal root ganglia (DRG) cells exhibited comparable changes, with 1% serum producing maximal increases in current magnitude (210%; p<.05), conductance (183%; p<.05), and rate of inactivation (71.4%; p<.001), in addition to inducing a hyperpolarizing shift in V50 (−9.402mV; p<.01). Future studies will focus on identifying this serum factor in order to evaluate its potential role in nociceptive signal modulation and cellular excitability. Supported in part by NIH grant R21DA034448 (SMT).