Identification of Ca_V channel types expressed in muscle afferent neurons

Abstract

Cardiovascular adjustments to exercise are partially mediated by group III/IV (small to medium) muscle afferents comprising the exercise pressor reflex (EPR). However, this reflex can be inappropriately activated in disease states (e.g., peripheral vascular disease), leading to increased risk of myocardial infarction. Here we investigate the voltage-dependent calcium (CaV) channels expressed in small to medium muscle afferent neurons as a first step toward determining their potential role in controlling the EPR. Using specific blockers and 5 mM Ba(2+) as the charge carrier, we found the major calcium channel types to be CaV2.2 (N-type) > CaV2.1 (P/Q-type) > CaV1.2 (L-type). Surprisingly, the CaV2.3 channel (R-type) blocker SNX482 was without effect. However, R-type currents are more prominent when recorded in Ca(2+) (Liang and Elmslie 2001). We reexamined the channel types using 10 mM Ca(2+) as the charge carrier, but results were similar to those in Ba(2+). SNX482 was without effect even though ∼27% of the current was blocker insensitive. Using multiple methods, we demonstrate that CaV2.3 channels are functionally expressed in muscle afferent neurons. Finally, ATP is an important modulator of the EPR, and we examined the effect on CaV currents. ATP reduced CaV current primarily via G protein βγ-mediated inhibition of CaV2.2 channels. We conclude that small to medium muscle afferent neurons primarily express CaV2.2 > CaV2.1 ≥ CaV2.3 > CaV1.2 channels. As with chronic pain, CaV2.2 channel blockers may be useful in controlling inappropriate activation of the EPR.

Inhibition of α9α10 nicotinic acetylcholine receptors prevents chemotherapy-induced neuropathic pain

“…L 4 and L 5 DRG neurons were isolated from Sprague-Dawley rats, enzymatically dissociated, plated onto polylysine-coated glass coverslips, and maintained overnight at 37°C in a 5% (vol/vol) CO 2 incubator in minimum essential media with 10% FBS and 1% penicillin/streptomycin (52). DRG neurons were recorded in a solution containing 145 mM N-methyl-D-glucosamine (NMG)•Cl, 5 mM BaCl 2 , 10 mM Na•Hepes, and 5 mM glucose, with pH 7.4 and osmolarity of 320 mOsm.…”

Cloning, synthesis, and characterization of αO-conotoxin GeXIVA, a potent α9α10 nicotinic acetylcholine receptor antagonist

“…The pipette solution contained 104 mM NMG·Cl, 14 mM creatine·PO 4 , 6 mM MgCl 2 , 10 mM NMG·Hepes, 5 mM Tris·ATP, 10 mM NMG 2 ·EGTA, and 0.3 Tris 2 ·GTP, with pH 7.4 and an osmolarity of 300 mOsm. Ionic currents were recorded and analyzed as previously described (19,58). Recordings were carried out at room temperature, and the holding potential was −80 mV.…”

“…S4). DRG calcium channels are composed of ∼50% N type (Ca V 2.2), ∼24% P/Q type (Ca V 2.1), and ∼13% L type (Ca V 1.2), with the remainder being R type (probably Ca V 2.3) (18)(19)(20).…”

Cardiac ion channels

“…35 Cav2.2 also plays a dominant role controlling the exercise pressor reflex through its expression in muscle afferents. 36 T-type channels differ from other VGCCs by their lack of accessory subunits and by their activation at more hyperpolarized voltages. T-type channels are expressed widely and play a key role in modulating excitability and burst firing in many cell types.…”