Functional expression of the organic cation/carnitine transporter 2 in rat astrocytes

In this study, we sought to identify the transporters that mediate the uptake of L-carnitine and acetyl-L-carnitine in cultured rat cortical astrocytes. L-[(3)H]carnitine and acetyl-L-[(3)H]carnitine uptake were both saturable, and mediated by a single Na(+)-dependent transport system. Uptake of both was inhibited by L-carnitine, D-carnitine, acetyl-L-carnitine and various organic cations. Acylcarnitines (acetyl-, butyryl-, hexanoyl-, octanoyl- and palmitoyl-L-carnitine) also interacted with L-[(3)H]carnitine and acetyl-L-[(3)H]carnitine transport. 2-Amino-2-norbornane carboxylic acid, a known inhibitor of amino acid transporter B(0,+) (ATB(0,+)), did not cause any significant inhibition. A highly significant correlation was found between the potencies of acylcarnitines in the inhibition of L-[(3)H]carnitine and acetyl-L-[(3)H]carnitine uptake and the acyl chain length of acylcarnitines. The expression of mRNA for organic cation/carnitine transporters (OCTNs), carnitine transporter 2 (CT2) and ATB(0,+) in astrocytes was investigated by reverse transcription (RT)-PCR. OCTN2 mRNA was expressed in astrocytes, whereas the expression of OCTN1, OCTN3 and CT2 mRNA could not be detected. ATB(0,+) mRNA was expressed at very low levels in astrocytes. Western blotting analysis indicated that anti-OCTN2 polyclonal antibody recognized a band of 70 kDa in both kidney and astrocyte preparations. OCTN2 immunoreactivity was detected in rat astrocytes by immunocytochemical staining. Inhibition of OCTN2 expression by RNA interference significantly inhibited L-[(3)H]carnitine and acetyl-L-[(3)H]carnitine uptake into astrocytes. These results suggest that OCTN2 is functionally expressed in rat astrocytes, and is responsible for L-carnitine and acetyl-L-carnitine uptake in these cells.