Expression of sperm Ca2+-activated K+ channels in Xenopus oocytes and their modulation by extracellular ATP.
Ionic fluxes across the sperm membrane have been shown to be important in the initiating process of sperm activation and gamete interaction; however, electrophysiological investigation of the ion channels involved has been precluded by the small size of the sperm, especially in mammalian species. In the present study sperm ion channels were expressed in Xenopus oocytes by injection of RNAs of spermatogenic cells isolated from the rat testes. The RNA-injected oocytes responded to ATP, a factor known to regulate sperm activation, with the activation of an outwardly rectifying whole-cell current which was dependent on K+ concentrations and inhibitable by K+ channel blockers, charybdotoxin (CTX) and tetraethylammonium (TEA). The ATP-induced current could be mimicked by a Ca2+ ionophore but suppressed by a Ca2+ chelator applied intracellularly, indicating a Ca2+ dependence of the current. Single-channel measurements on RNA-injected oocytes revealed channels of large conductance which could be blocked by CTX and TEA. Co-injection of germ cell RNAs with the antisense RNA for a mouse gene encoding slowpoke 'Maxi' Ca2+-activated K+ channels resulted in significant reduction of the ATP- and ionomycin-induced current. The expression of the 'Maxi' Ca2+-activated K+ channels in sperm collected from the rat epididymis was also confirmed by Western blot analysis. These results suggest that sperm possess Ca2+-activated K+ channels which may be involved in the process of sperm activation.
Chan
H C
HC
Department of Physiology, Faculty of Medicine, The Chinese University of Hong Kong, Shatin. hsiaocchan@cuhk.edu.hk
Wu
W L
WL
Sun
Y P
YP
Leung
P S
PS
Wong
T P
TP
Chung
Y W
YW
So
S C
SC
Zhou
T S
TS
Yan
Y C
YC
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D002118:Calcium / Q000378:metabolism; D002772:Cholera Toxin / Q000494:pharmacology; D017259:Dimaprit / Q000031:analogs & derivatives* / Q000378:metabolism; D005260:Female; D015759:Ionomycin / Q000494:pharmacology; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008297:Male; D008322:Mammals; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D008969:Molecular Sequence Data; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology; D011188:Potassium / Q000494:pharmacology / Q000502:physiology; D015221:Potassium Channels / Q000032:analysis / Q000096:biosynthesis / Q000502:physiology*; D024681:Potassium Channels, Calcium-Activated; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000096:biosynthesis; D012741:Sexual Maturation; D013094:Spermatozoa / Q000378:metabolism / Q000502:physiology*; D014982:Xenopus laevis
D000906:Antibodies; C494842:Kcnma1 protein, rat; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051036:Large-Conductance Calcium-Activated Potassium Channels; D010446:Peptide Fragments; D015221:Potassium Channels; D024681:Potassium Channels, Calcium-Activated; D011994:Recombinant Proteins; D015759:Ionomycin; C071989:APT; D002772:Cholera Toxin; D011188:Potassium; D002118:Calcium; D017259:Dimaprit