Effect of a prolonged glutamate challenge on plasmalemmal calcium permeability in mammalian central neurones. Mn2+ as a tool to study calcium influx pathways.
The rate of Mn(2+)-induced fluorescence quenching (RFQ) was used as a relative measure of plasma membrane Ca2+ permeability (PCa) in fura-2-loaded cultured hippocampal neurons and cerebellar granule cells during and after protracted (15-30 min) glutamate (GLU) treatment. Some limitations of this method were evaluated using a kinetic model of a competitive binding of Mn2+ and Ca2+ to fura-2 in the cell. In parallel experiment a contribution of Ca2+ influx to the cytoplasmic Ca2+ ([Ca2+]i) was repeatedly examined during and following a prolonged GLU challenge by short-duration "low-Ca2+ trials" (50 microM EGTA) and by measurements of 45Ca2+ uptake. Experiments failed to reveal a putative persistent increase in PCa that earlier was thought to underlie Ca2+ overload of the neuron caused by its toxic GLU treatment. By contrast, a sustained increase of [Ca2+]i was found to be associated with a progressive decrease in PCa and Ca2+ influx both in the period of GLU application and after its termination. These findings give new evidence in favour of the hypothesis that the GLU-induced Ca2+ overload of the neuron mainly from an impairment of its Ca2+ extrusion systems.
Khodorov
B I
BI
Institute of General Pathology and Pathophysiology, Russian Academy of Medical Sciences, Moscow, Russia. rans@rans.msk.ru
Fayuk
D A
DA
Koshelev
S G
SG
Vergun
O V
OV
Pinelis
V G
VG
Vinskaya
N P
NP
Storozhevykh
T P
TP
Arsenyeva
E N
EN
Khaspekov
L G
LG
Lyzhin
A P
AP
Isaev
N
N
Victorov
I V
IV
Dubinsky
J M
JM
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects*; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D005453:Fluorescence; D018698:Glutamic Acid / Q000633:toxicity*; D007476:Ionophores / Q000494:pharmacology; D008345:Manganese; D016202:N-Methylaspartate / Q000633:toxicity; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D009532:Nickel / Q000633:toxicity; D051381:Rats; D017208:Rats, Wistar
D015220:Calcium Channels; D002614:Chelating Agents; D007476:Ionophores; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D018698:Glutamic Acid; D008345:Manganese; D004533:Egtazic Acid; D016202:N-Methylaspartate; D009532:Nickel; D002118:Calcium