Voltage-Dependent Ion Channels in CAD Cells: A Catecholaminergic Neuronal Line That Exhibits Inducible Differentiation

Abstract

Cell lines derived from tumors engineered in the CNS offer promise as models of specific neuronal cell types. CAD cells are an unusual subclone of a murine cell line derived from tyrosine hydroxylase (TH) driven tumorigenesis, which undergoes reversible morphological differentiation on serum deprivation. Using single-cell electrophysiology we have examined the properties of ion channels expressed in CAD cells. Despite relatively low resting potentials, CAD cells can be induced to fire robust action potentials when mildly artificially hyperpolarized. Correspondingly, voltage-dependent sodium and potassium currents were elicited under voltage clamp. Sodium currents are TTX sensitive and exhibit conventional activation and inactivation properties. The potassium currents reflected two pharmacologically distinguishable populations of delayed rectifier type channels while no transient A-type channels were observed. Using barium as a charge carrier, we observed an inactivating current that was completely blocked by nimodipine and thus associated with L-type calcium channels. On differentiation, three changes in functional channel expression occurred; a 4-fold decrease in sodium current density, a 1.5-fold increase in potassium current density, and the induction of a small noninactivating barium current component. The neuronal morphology, excitability properties, and changes in channel function with differentiation make CAD cells an attractive model for study of catecholaminergic neurons.

Proteasome inhibitors promote the sequestration of PrPSc into aggresomes within the cytosol of prion-infected CAD neuronal cells

“…The CAD cell line was used here as the main model system. These cells derive from mouse brain neurons, and on serum deprivation exhibit a differentiated phenotype with major ultrastructural and electrophysiological features of CNS neuronal cells (Qi et al, 1997;Wang & Oxford, 2000). We found that, whilst CAD cells did accumulate abnormal PrP following treatment with proteasome inhibitors, the formation of aggresomes containing aggregated PrP with true PrP Sc characteristics was only a feature of prion-infected cultures.…”

In Silico Docking and Electrophysiological Characterization of Lacosamide Binding Sites on Collapsin Response Mediator Protein-2 Identifies a Pocket Important in Modulating Sodium Channel Slow Inactivation

“…CAD Cells-Catecholamine A-differentiated (CAD) cells were grown at 37°C and in 5% CO 2 (Sarstedt, Newton, NC) in Ham's F-12/Eagle's minimum essential media (Amersham Biosciences), supplemented with 8% fetal bovine serum (Sigma) and 1% penicillin/streptomycin (100% stocks, 10,000 units/ml penicillin G sodium, and 10,000 g/ml streptomycin sulfate) (25). Cells were passaged every 6 -7 days at a 1:25 dilution.…”

“…Using the whole cell patch clamp configuration, the effects of LCM on VGSCs in the presence of overexpressed CRMP-2 were examined. CAD cells were chosen, because they 1) provide a background easily amenable to genetic manipulation, 2) are of neuronal origin, and 3) express endogenous tetrodotoxin-sensitive Na ϩ currents (25). Current-voltage relationships in control EGFP or CRMP-2-EGFP-expressing CAD cells were examined by the application of 15-ms step depolarizations ranging from Ϫ70 mV to ϩ80 mV (in ϩ10-mV increments) from a holding potential of Ϫ80 mV (Fig.…”

Myo10 in brain: developmental regulation, identification of a headless isoform and dynamics in neurons

“…CAD cells were cultured in HAM-F12 with 8% fetal bovine serum (FBS) and 1% penicillin/streptomycin as previously described (Wang and Oxford, 2000). Undifferentiated CAD cells were transiently transfected at 60-80% confluence using Polyfect (Qiagen) for 12 hours following to the manufacturer's instructions for HeLa cells.…”