Different voltage-gated sodium currents are expressed by human neuroblastoma NB69 cells when cultured in defined serum-free and in astroglial-conditioned media.
Voltage-gated Na+ currents (INa) were analysed with the whole-cell patch-clamp technique in human neuroblastoma NB69 cells plated in serum-free "defined" medium (DM) or in "astroglial-conditioned" medium (CM). Cells survived in both media and expressed the microtubule associated protein 1A, indicating neuron-like differentiation. Two INa types with different time-, voltage-dependent properties and tetrodotoxin (TTX) sensitivities were expressed in DM and CM. The INa in DM-plated cells was present from day 4 and its surface density increased from 11 pA/pF (days 5-7) to 68 pA/pF (days 15-30). The underlying conductance (GNa) half-activated (V0A) at -24 mV. INa inactivation was fitted by single exponentials with 7.5 ms time constant (th) at the -35 mV half-inactivation voltage (V0I). INa was not affected by 10 nM, was reduced (65%) by 100 nM, and not completely abolished (92%) by 300 nM tetrodotoxin (TTX). The INa of CM-plated cells appeared at day 3-4 and its surface density increased from 14 pA/pF (days 3-6) to 28 pA/pF (days 11-14). The GNa V0A was -29 mV and inactivation was fitted by single exponentials with 2.6 ms that the -58 mV V0I. This INa was reduced (55%) by 10 nM and totally abolished by 100 nM tetrodotoxin (TTX). In conclusion, NB69 cells displayed a slow, "TTX-resistant," or a fast, "TTX-sensitive" INa in DM and CM, respectively, suggesting that the CM contained diffusible trophic factors of astroglial origin that induced the expression of a different Na+ channel type. About half of the CM- and DM-plated cells also displayed a persistent Na+ current (INaP).
Urbano
F J
FJ
Instituto Cajal, CSIC, Madrid, Spain.
Sierra
F
F
Colombo
J A
JA
Velasco
J M
JM
Buño
W
W
D000200:Action Potentials / Q000502:physiology; D001253:Astrocytes / Q000502:physiology*; D017077:Culture Media, Conditioned / Q000494:pharmacology; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects* / Q000502:physiology; D009447:Neuroblastoma; D018408:Patch-Clamp Techniques; D015222:Sodium Channels / Q000502:physiology*; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000187:drug effects / Q000502:physiology
D017077:Culture Media, Conditioned; D016895:Culture Media, Serum-Free; D015222:Sodium Channels