Gap junctions do not underlie changes in whole-cell conductance in anoxic turtle brain.
An acute reduction in cell membrane permeability could provide an effective strategy to prolong anoxic survival. A previous study has shown that in the western painted turtle whole-cell neuronal conductance (G(w)) decreases during anoxia, which may be mediated by the activation of adenosine A(1) receptors and calcium. Reduction in G(w) is thought to be the result of ion channel closure, but closure of gap junctions could also be responsible for this phenomenon. In our study, antibody staining of connexin 32 and 43 (Cx32 and Cx43) suggested the presence of gap junctional components in the turtle cortex. To examine if gap junctions were involved in the previously measured anoxic decrease in G(w), neuronal connectivity was assessed through the measurement of whole-cell capacitance (C(w)). Turtle cortical sheets were perfused with normoxic (95%O(2)/5%CO(2)), anoxic (95%N(2)/5%CO(2)), high calcium (4 mM) and adenosine (200 microm) artificial cerebral spinal fluid (aCSF). No significant change in C(w) was observed under any of the above conditions. However, during hypo-osmotic aCSF perfusion C(w) decreased significantly, with the lowest value of 50+/-10.4 pF (P<0.05) occurring at 30 min. To visualize changes in gap junction permeability lucifer yellow was loaded into turtle neurons during normoxic, anoxic, 0 calcium, hypo-osmotic, cold shock, (+)-isoproterenol, nitric oxide donor S-nitoso-acetyl penicillamine, and 8-bromo-guanosine 3',5'-cyclic monophosphate aCSF perfusion. Dye propagation was only observed in 3 of 20 cold shock experiments (4 degrees C). We conclude that gap junctions are not involved in the acute reduction in G(w) previously observed during anoxia and that our results support the hypothesis that ion channel arrest is involved.
Shin
Damian Seung-ho
DS
Department of Zoology, University of Toronto, 25 Harbord Street, Toronto, Ont, Canada M5S 3G5.
Ghai
Himesh
H
Cain
Sean Wendell
SW
Buck
Leslie Thomas
LT
D000818:Animals; D002450:Cell Communication / Q000502:physiology; D018031:Connexin 43 / Q000032:analysis; D017630:Connexins / Q000032:analysis; D025802:Electric Capacitance; D004553:Electric Conductivity; D005260:Female; D005456:Fluorescent Dyes; D017629:Gap Junctions / Q000737:chemistry / Q000502:physiology*; D020925:Hypoxia-Ischemia, Brain / Q000503:physiopathology*; D007546:Isoquinolines; D009474:Neurons / Q000737:chemistry / Q000502:physiology*; D018408:Patch-Clamp Techniques; D014426:Turtles / Q000502:physiology*; D000097002:Gap Junction beta-1 Protein
D018031:Connexin 43; D017630:Connexins; D005456:Fluorescent Dyes; D007546:Isoquinolines; D000097002:Gap Junction beta-1 Protein; C017475:lucifer yellow