Molecular and functional identification of large neutral amino acid transporters LAT1 and LAT2 and their pharmacological relevance at the blood-brain barrier.

Abstract

We present here the evidence of molecular and functional expression of LAT1 and LAT2, subunits of the large neutral amino acid transporter system L, in cultured brain capillary endothelial cells of the rat. By means of the RT-PCR method, transcripts of LAT1, LAT2 and heavy chain of 4F2 antigen (4F2hc) were detected in rat primary cultured brain capillary endothelial cells (BCECs) and immortalized subline, RBEC1. The uptake properties of RBEC1, such as [3H]leucine and L-[3H]DOPA uptake, were similar to those of primary cultured BCECs. So, RBEC1 may retain almost native properties of the large neutral amino acid uptake activities. [3H]Leucine uptake by RBEC1 showed two saturable components and the Km values of the high- and low-affinity components were 8.92+/-3.18 and 119+/-45 microM, respectively. The Km value of the high-affinity component agreed well with that of LAT1 and the amino acid transport selectivity of RBEC1 was similar to that of LAT1. Therefore, it is suggested that LAT1 is important at the blood-brain barrier of rats. Additionally, the Km value of the low-affinity component was similar to that of LAT2. These observations indicate that LAT1 and LAT2 are involved as transporters for large neutral amino acids at the blood-brain barrier. Additionally, we concluded that RBEC1 is useful as an in-vitro model for evaluation of the pharmacological relevance of system L at the blood-brain barrier.