Taurine transporter in primary cultured neonatal rat heart cells: a comparison between cardiac myocytes and nonmyocytes.
In the present study, we examined the characteristics of the taurine transporter and the intracellular taurine content in cultured neonatal heart cells. Primary cultures of cardiac myocytes and cardiac fibroblasts (nonmyocytes) were prepared from 1-day-old Wistar rats. The parameters examined were: (a) intracellular taurine content by the HPLC method, (b) the expression levels of taurine transporter mRNA and protein using northern and western blot analysis, and (c) transporter activity determined by the uptake of 3H-labeled taurine. The taurine content of myocytes was significantly higher (3-fold) than that of nonmyocytes. Taurine transporter mRNA was strongly expressed in both myocytes and nonmyocytes, whereas the magnitude [normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene expression] of the transporter mRNA expressed in myocytes was lower than that in nonmyocytes. The expression level of transporter protein in myocytes was also lower than that of nonmyocytes. Uptake of radiolabeled taurine into monolayer cultures of heart cells was stimulated markedly by the presence of Na+ in the medium, whereas this uptake was almost abolished in the absence of Na+. The Na+/taurine stoichiometry was 2:1 for both myocytes and nonmyocytes. Kinetic analysis showed that a single saturable system was involved in taurine uptake into both cell types. In myocytes, the apparent K(m) and V(max) values for the transporter were 20.7+/-0.5 microM and 1.07+/-0.01 nmol/10(6)cells/30 min, respectively. Similarly, those of nonmyocytes were 20.3+/-0.7 microM and 0.42+/-0.01 nmol/10(6)cells/30 min. These findings indicated that both myocytes and nonmyocytes expressed an identical taurine transporter with a Michaelis-Menten constant of 20-21 microM and that a higher taurine content in myocytes may be associated with a higher V(max).
Takahashi
Kyoko
K
Department of Clinical Evaluation of Medicines and Therapeutics, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
Azuma
Mistuhiro
M
Yamada
Tomoko
T
Ohyabu
Yuko
Y
Takahashi
Koichi
K
Schaffer
Stephen W
SW
Azuma
Junichi
J
D000818:Animals; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002478:Cells, Cultured; D008562:Membrane Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D026901:Membrane Transport Proteins; D032342:Muscle Cells / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D013654:Taurine / Q000378:metabolism*; D000099247:Taurine Transporters
D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D013654:Taurine; D000099247:Taurine Transporters