Type two voltage gated calcium (Ca V 2) channels are the primary mediators of neurotransmission at neuronal presynapses, but their function at neural soma is also important in regulating excitability. 1 Mechanisms that regulate Ca V 2 channel expression at synapses have been studied extensively, which motivated us to perform similar studies in the soma. Rat sympathetic neurons from the superior cervical ganglion (SCG) natively express Ca V 2.2 and Ca V 2.3. 2 We noted previously that heterologous expression of Ca V 2.1 but not Ca V 2.2 results in increased calcium current in SCG neurons. 3 In the present study, we extended these observations to show that both Ca V 2.1 and Ca V 2.3 expression resulted in increased calcium currents while Ca V 2.2 expression did not. Further, Ca V 2.1 could displace native Ca V 2.2 channels, but Ca V 2.3 expression could not. Heterologous expression of the individual accessory subunits a 2 d-1, a 2 d-2, a 2 d-3, or b4 alone failed to increase current density, suggesting that the calcium current ceiling when Ca V 2.2 was over-expressed was not due to lack of these subunits. Interestingly, introduction of recombinant a2d subunits produced surprising effects on displacement of native Ca V 2.2 by recombinant channels. Both a 2 d-1 and a 2 d-2 seemed to promote Ca V 2.2 displacement by recombinant channel expression, while a 2 d-3 appeared to protect Ca V 2.2 from displacement. Thus, we observe a selective prioritization of Ca V channel functional expression in neurons by specific a 2 d subunits. These data highlight a new function for a 2 d subtypes that could shed light on subtype selectivity of Ca V 2 membrane expression.

Long QT type 3 (LQT3) is a common form of LQT syndrome caused by mutations in the cardiac sodium channel. These mutations lead to an increase in the cardiac sodium current during the late phases of the cardiac action potential (AP), leading to cardiac arrhythmias. We attempted to predict mutation-specific cardiac event risk for LQT3 mutations using a combination of in vitro and in silico risk assessment. For that, we first characterized, using patch clamp, eight common mutations associated with LQT3. Using our data as an input to a cardiomyocyte computer model, we predicted action potential duration (APD) and early after-depolarization (EAD) susceptibility for these mutants, at different heart rates. Our risk predictions correlated well to the cardiac event risk observed in patients with these mutations. Second, we measured the effect of the late sodium blocker ranolazine for each of the mutants tested. We used the model to predict drug treatment efficacy. Interestingly, ranolazine showed mutation-specific effects, with some mutations showing strongly shortened APDs and decreased predicted risk, while little effect was observed in other mutants. Most importantly, for one of the mutations assessed, risk was predicted to be increased, despite shortening of APD. Thus, our results suggest that drug treatment efficacy is mutation-specific. In summary, our results show that mutation-specific in silico models may be an important new tool for clinical risk assessment in LQT3.

Voltage gated calcium channels (Cav) are comprised of several different subunits necessary to achieve proper function. Some of the subunits (β, α2‐γ) contribute to not only channel properties, but to trafficking. In heterologous expression systems that lack endogenous Cav channels, pore‐forming subunits must be co‐expressed with accessory subunits for channel localization to the plasma membrane and for proper function. In some neurons expression of an exogenous pore, without co‐expression of its respective subunits, can result in a functional channel. We investigate the channel expression in the cell body of rat superior cervical ganglion neurons (SCG) using whole cell patch clamping. SCG neurons express mainly type 2 Cav channels. Interestingly, when the cDNA encoding Cav pore‐forming subunits are injected intranuclearly into SCG neurons, large increases in current density corresponding to recombinant Cav channel expression are observed in a subunit specific manner. These data suggest that pre‐formed, sub‐unit specific “slots” exist in SCG neurons to allow for the expression, trafficking and function of certain Cav channel subunits, but not others. Using this approach, we observe significant increases in calcium current density following the expression of Cav 2.1, mCav2.3, Cav 1.3 subunits, but no increase in current density when Cav 2.2 or Cav 1.2 are injected. RO1 GM101023