Adult neural precursor cells form connexin-dependent networks that improve their survival.
Establishment of cellular networks and calcium homeostasis are essential for embryonic stem cell proliferation and differentiation. We also hypothesized that adult neural progenitor cells form functional cellular networks relevant for their development. We isolated neuronal progenitor cells from the subventricular zone of 5-week-old mice to investigate the role of gap junctions, calcium homeostasis, and cellular networks in cell differentiation and survival. Western blotting and reverse transcription-PCR showed that the cells expressed the gap junction components connexin 26, 36, 43, and 45, and that expression of connexin 43 increased in early (8 days) differentiated cells. Transmission electron microscopy and immunocytochemistry also indicated that gap junctions were present. Scrape-loading experiments showed dye transfer between cells that could be prevented by gapjunction blockers; thus, functional intercellular gap junctions had been established. However, dye transfer was four times stronger in differentiated cultures, correlating with the increased connexin 43 expression. During time-lapse calcium imaging, both differentiated and undifferentiated cultures showed spontaneous calcium activity that was reduced by gap junction blockers. Cross-correlation analysis of the calcium recordings showed that the cells were interconnected through gap junctions and that the early-differentiated cells were organized in small-world networks. Gap junction blockers did not affect proliferation and differentiation, but resulted in twice as many apoptotic cells. mRNAi knockdown of connexin 43 also doubled the number of apoptotic cells. We conclude that adult neural progenitor cells form networks in vitro that are strengthened during early differentiation by increased expression of connexin 43. The networks are functional and improve cell survival.
aDepartment of Women's and Children's Health, Karolinska Institutet, Stockholm, Sweden bLaboratory of Experimental Neurology cLaboratory of Histology, Neuroanatomy and Neuropathology, Free University of Brussels (ULB), Brussels, Belgium.
Ajaya
A
Ravella
Thomas
T
Ringstedt
Jean-Pierre
JP
Brion
Massimo
M
Pandolfo
Eric
E
Herlenius
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling; D002450:Cell Communication / Q000502:physiology*; D002478:Cells, Cultured; D002552:Cerebral Ventricles / Q000166:cytology; D017630:Connexins / Q000235:genetics / Q000378:metabolism*; D017629:Gap Junctions / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D020287:In Situ Nick-End Labeling; D051379:Mice; D008822:Mice, Transgenic; D008854:Microscopy, Electron; D058953:Neural Stem Cells / Q000378:metabolism*; D034622:RNA Interference / Q000502:physiology; D012333:RNA, Messenger; D013223:Statistics as Topic
D017630:Connexins; D012333:RNA, Messenger; D049452:Green Fluorescent Proteins; D002118:Calcium