Subunit‐dependent cadmium and nickel inhibition of acid‐sensing ion channels

Abstract

Acid-sensing ion channels (ASIC) are ligand-gated cation channels that are highly expressed in peripheral sensory and central neurons. ASIC are transiently activated by decreases in extracellular pH and are thought to play important roles in sensory perception, neuronal transmission, and excitability, and in the pathology of neurological conditions, such as brain ischemia. We demonstrate here that the heavy metals Ni 21 and Cd 21 dose-dependently inhibit ASIC currents in hippocampus CA1 neurons and in Chinese hamster ovary (CHO) cells heterologously expressing these channels. rapidly inhibited homomeric ASIC1a and heteromeric ASIC1a/2a channels without affecting ASIC1b, 2a, and ASIC3 homomeric channels and ASIC1a/3 and 2a/3 heteromeric channels. In contrast, external Cd 21 (1 mM) inhibited ASIC2a and ASIC3 homomeric channels and ASIC1a/2a, 1a/3, and 2a/3 heteromeric channels but not ASIC1a homomeric channels. The acid-sensing current in isolated rat hippocampus CA1 neurons, thought to be carried primarily by ASIC1a and 1a/2a, was inhibited by 1 mM Ni 21 . The current study identifies ASIC as a novel target for the neurotoxic heavy metals Cd 21 and Ni 2þ .

Inhibition of Neuronal Degenerin/Epithelial Na+ Channels by the Multiple Sclerosis Drug 4-Aminopyridine

“…The expression vector encoding YFP-tagged rASIC1b was a gift from Drs. D. J. Benos and C. Fuller (University of Alabama-Birmingham, AL) and has been described previously (34,35). The expression vector encoding constitutively active rat BLINaC harboring the A443T mutation was a gift from Dr. E. Lingueglia (Institut de Pharmacologie Moleculaire et Cellulaire, Valbonne, France) and has been described previously (24).…”

“…Cell Culture-Primary CA1 hippocampal neurons were obtained from newborn (P1-P3) Sprague-Dawley rats and C57BL/6 mice using standard procedures (34,36,37). All animal use and welfare adhered to the National Institutes of Health Guide for the Care and Use of Laboratory Animals following a protocol reviewed and approved by the Institutional Laboratory Animal Care and Use Committee of the University of Texas Health Science Center at San Antonio.…”

“…Expression of Recombinant ASIC and BLINaC in CHO Cells-CHO cells were transfected with plasmids encoding BLINaC or ASIC subunits using Polyfect reagent (Qiagen) following the manufacturer's protocol as described previously (34). For expression of homomeric channels, cells were transfected with 1 g of rASIC1a, rASIC1b, rASIC2a, rACIS3, or rBLINaC cDNA/35-mm 2 dish.…”

“…Patch Clamp Electrophysiology-Acid-sensing macroscopic currents conducted by ASIC in hippocampal CA1 neurons were measured in the whole cell patch clamp configuration under voltage clamp conditions using standard methods (34). In brief, neurons were clamped to Ϫ60 mV, and currents were activated by rapid solution exchange from control (7.4) to acidic (5.0) pH.…”

“…Whole cell macroscopic current recordings of ASIC expressed in CHO cells were performed under voltage-clamp conditions (V m ϭ Ϫ10 mV) using standard methods (34). In these experiments, ASIC currents were evoked by rapid solution exchange from control (7.6) to acidic (5.0) pH.…”