Differentiation of ionic currents in CNS progenitor cells: dependence upon substrate attachment and epidermal growth factor.
Multipotential progenitor cells grown from central nervous system (CNS) tissues in defined media supplemented with epidermal growth factor (EGF), when attached to a suitable substratum, differentiate to express neural and glial histochemical markers and morphologies. To assess the functional characteristics of such cells, expression of voltage-gated Na+ and K+ currents (INa, IK) was studied by whole-cell patch clamp methods in progenitors raised from postnatal rat forebrain. Undifferentiated cells were acutely dissociated from proliferative "spheres," and differentiated cells were studied 1-25 days after plating spheres onto polylysine/laminin-treated coverslips. INa and IK were detected together in 58%, INa alone in 11%, and IK alone in 19% of differentiated cells recorded with K(+)-containing pipettes. With internal Cs+ (to isolate INa), INa up to 45 pA/pF was observed in some cells within 1 day after plating. I Na ranged up to 150 pA/pF subsequently. Overall, 84% of cells expressed I Na, with an average of 38 pA/pF. INa had fast kinetics, as in neurons, but steadystate inactivation curves were strongly negative, resembling those of glial INa. Inward tail currents sensitive to [K+]out were observed upon repolarization after the 10-ms test pulse with internal Cs+, indicating the expression of K+ channels in 82% of cells. In contrast to the substantial currents observed in differentiating cells, little or no INa or Ik-tail currents were detected in recordings from cells acutely dissociated from spheres. Thus, in the presence of EGF, ionic currents develop early during differentiation induced by attachment to an appropriate substratum. Cells switched from EGF to basic fibroblast growth factor (bFGF) when plated onto coverslips showed greatly reduced proliferation and developed less neuron-like morphologies than cells plated in the presence of EGF. INa was observed in only 53% of bFGF-treated cells, with an average of 9 pA/pF. Thus, in contrast to reports that bFGF promotes neuronal differentiation in some CNS progenitor populations, our EGF-generated postnatal rat CNS progenitors do not develop neuronal characteristics when switched to medium containing bFGF. Thus, differentiated CNS progenitors can express a mix of neuronal and glial molecular, morphological, and electrophysiological properties that can be modified by culture conditions.
Feldman
D H
DH
Department of Neuroscience, College of Medicine, University of Florida Brain Institute, Gainesville 32610, USA.
Thinschmidt
J S
JS
Peel
A L
AL
Papke
R L
RL
Reier
P J
PJ
D000818:Animals; D000831:Animals, Newborn; D055592:Biophysical Phenomena; D001703:Biophysics; D001923:Brain Chemistry / Q000502:physiology; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002454:Cell Differentiation; D002478:Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D002490:Central Nervous System / Q000166:cytology*; D003470:Culture Media / Q000494:pharmacology; D004594:Electrophysiology; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005069:Evaluation Studies as Topic; D015640:Ion Channel Gating / Q000502:physiology; D007473:Ion Channels / Q000502:physiology*; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology; D016548:Prosencephalon / Q000166:cytology; D051381:Rats; D015222:Sodium Channels / Q000187:drug effects / Q000502:physiology; D013234:Stem Cells / Q000737:chemistry* / Q000166:cytology / Q000502:physiology
D003470:Culture Media; D007473:Ion Channels; D015221:Potassium Channels; D015222:Sodium Channels; D004815:Epidermal Growth Factor