Overexpression of potassium channel RNA: in vivo development rescues neurons from suppression of morphological differentiation in vitro.
Neuronal differentiation often proceeds differently in vitro than it does in vivo. Previous work demonstrated that overexpression of potassium channel RNA reduces the number of morphologically identifiable neurons that appear in cultures prepared from neural plate stage (17-1/2 hr) embryos (Jones and Ribera, 1994). Here, we report that morphological differentiation of neurons in situ is only slightly affected by overexpression of potassium channels. Endogenous factors appear to compensate for the effect of channel overexpression. Consistent with this view, when cultures are prepared from older neural tube embryos (22-24 hr), more neurons containing excess potassium channel RNA differentiate morphologically in vitro. Exposure in situ to a rapid intracellular calcium chelator, but not to tetrodotoxin, omega-conotoxin or a slow calcium chelator, prevents the compensation provided by extended development in vivo. Typically, RNA overexpression is limited to half of the embryo in order to provide an internal control. However, when potassium channel RNA is overexpressed throughout the embryo, few neurons differentiate morphologically in vitro, even if cultures are prepared from older neural tube embryos. Thus, recovery is possible if a minimum of 5 hr of further development in vivo is allowed under conditions in which rapid elevations of intracellular calcium are permitted and half of the nervous system has normal levels of potassium channel RNA. These results suggest that different or additional mechanisms operate in situ than in vitro to promote morphological differentiation of neurons.
Jones
S M
SM
Department of Physiology, University of Colorado Health Sciences Center, Denver 80262, USA.
Hofmann
A D
AD
Lieber
J L
JL
Ribera
A B
AB
D000818:Animals; D001757:Blastomeres / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004625:Embryo, Nonmammalian; D015870:Gene Expression / Q000187:drug effects; D006321:Heart / Q000187:drug effects / Q000502:physiology; D007150:Immunohistochemistry; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology; D015221:Potassium Channels / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000096:biosynthesis*; D013779:Tetrodotoxin / Q000494:pharmacology; D014981:Xenopus; D020960:omega-Conotoxins
D002121:Calcium Channel Blockers; D010455:Peptides; D015221:Potassium Channels; D012333:RNA, Messenger; D020960:omega-Conotoxins; C053423:Conus magus toxin; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D013779:Tetrodotoxin; D004533:Egtazic Acid