Reincorporated plasma membrane Ca2+-ATPase can mediate B-Type Ca2+ channels observed in native membrane of human red blood cells

Autor: Pinet, C., Antoine, S., Filoteo, A.G., Penniston, J.T., Coulombe, A.
Přispěvatelé: CNRS UMR 8078
Jazyk: angličtina
Rok vydání: 2002
Předmět:
Erythrocytes
Patch-Clamp Techniques
Chlorpromazine
Proteolipids
[SDV]Life Sciences [q-bio]
MESH: Electric Conductivity
Calcium-Transporting ATPases
Membrane Potentials
MESH: Calcium-Transporting ATPases
MESH: Dose-Response Relationship
Drug
Plasma Membrane Calcium-Transporting ATPases
MESH: Cation Transport Proteins
Adenosine Triphosphate
Calmodulin
MESH: Adenosine Triphosphate
MESH: Patch-Clamp Techniques
Humans
MESH: Membrane Potentials
Cation Transport Proteins
MESH: Humans
Dose-Response Relationship
Drug
MESH: Erythrocytes
Cell Membrane
Electric Conductivity
MESH: Chlorpromazine
MESH: Calmodulin
MESH: Vanadates
MESH: Calcium Channels
Eosine Yellowish-(YS)
MESH: Eosine Yellowish-(YS)
MESH: Plasma Membrane Calcium-Transporting ATPases
Calcium Channels
Vanadates
MESH: Proteolipids
MESH: Cell Membrane
Zdroj: Journal of Membrane Biology
Journal of Membrane Biology, Springer Verlag (Germany), 2002, 187 (3), pp.185-201. ⟨10.1007/s00232-001-0163-5⟩
ISSN: 0022-2631
1432-1424
DOI: 10.1007/s00232-001-0163-5⟩
Popis: International audience; Recently, we reported indirect evidence that plasma membrane Ca2+-ATPase (PMCA) can mediate B-type Ca2+ channels of cardiac myocytes. In the present study, in order to bring more direct evidence, purified PMCA from human red blood cells (RBC) was reconstituted into giant azolectin liposomes amenable to the patch-clamp technique. Purified RBC PMCA was used because it is available pure in larger quantity than cardiac PMCA. The presence of B-type Ca2+ channels was first investigated in native membranes of human RBC. They were detected and share the characteristics of cardiac myocytes. They spontaneously appeared in scarce short bursts of activity, they were activated by chlorpromazine (CPZ) with an EC50 of 149 mmole/l or 1 mmole/l vanadate, and then switched off by 10 mmole/l eosin or dose-dependently blocked by 1-5 mmole/l ATP. Independent of membrane potential, the channel gating exhibited complex patterns of many conductance levels, with three most often observed conductance levels of 22, 47 and 80 pS. The activation by vanadate suggests that these channels could play a role in the influx of extracellular Ca2+ involved in the vanadate-induced Gardos effect. In PMCA-reconstituted proteoliposomes, nearly half of the ATPase activity was retained and clear "channel-like" openings of Ba2+- or Ca2+-conducting channels were detected. Channel activity could be spontaneously present, lasting the patch lifetime or, when previously quiescent, activity could be induced by application of 50 mmole/l CPZ only in presence of 25 U/ml calmodulin (CaM), or by application of 1 mmole/l vanadate alone. Eosin (10 mmole/l) and ATP (5 mmole/l) significantly reduced spontaneous activity. Channel gating characteristics were similar to those of RBC, with main conductance levels of 21, 40 and 72 pS. The lack of direct activation by CPZ alone might be attributed to a purification-induced modification or absence of unidentified regulatory component(s) of PMCA. Despite a few differences in results between RBC and reincorporated PMCA, most probably attributable to the decrease in ATPase activity following the procedure of reincorporation, the present experimental conditions appear to reveal a channel-mode of the PMCA that shares many similarities with the B-type Ca2+ channel.
Databáze: OpenAIRE
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