Showing papers by "Masato Inazu published in 2019"
TL;DR: CTL1- and CTL2-mediated choline transport to the brain through the blood–brain barrier plays an essential role in various functions of the central nervous system by acting as the rate-limiting step of cholinergic neuronal activity.
Abstract: Cholinergic neurons in the central nervous system play a vital role in higher brain functions, such as learning and memory. Choline is essential for the synthesis of the neurotransmitter acetylcholine by cholinergic neurons. The synthesis and metabolism of acetylcholine are important mechanisms for regulating neuronal activity. Choline is a positively charged quaternary ammonium compound that requires transporters to pass through the plasma membrane. Currently, there are three groups of choline transporters with different characteristics, such as affinity for choline, tissue distribution, and sodium dependence. They include (I) polyspecific organic cation transporters (OCT1-3: SLC22A1-3) with a low affinity for choline, (II) high-affinity choline transporter 1 (CHT1: SLC5A7), and (III) choline transporter-like proteins (CTL1-5: SLC44A1-5). Brain microvascular endothelial cells, which comprise part of the blood–brain barrier, take up extracellular choline via intermediate-affinity choline transporter-like protein 1 (CTL1) and low-affinity CTL2 transporters. CTL2 is responsible for excreting a high concentration of choline taken up by the brain microvascular endothelial cells on the brain side of the blood–brain barrier. CTL2 is also highly expressed in mitochondria and may be involved in the oxidative pathway of choline metabolism. Therefore, CTL1- and CTL2-mediated choline transport to the brain through the blood–brain barrier plays an essential role in various functions of the central nervous system by acting as the rate-limiting step of cholinergic neuronal activity.
38 citations
TL;DR: It is demonstrated for the first time that TRPM7 was functionally expressed in both freshly isolated white adipocytes and in 3T3‐L1 adipocyte cell line by whole‐cell patch‐clamp recordings, and plays a role as a Ca2+ influx pathway in adipocytes.
Abstract: In adipocytes, intracellular Ca2+ and Mg2+ modulates physiological functions, such as insulin action and the secretion of adipokines. TRPM7 is a Ca2+ /Mg2+ -permeable non-selective cation channel. TRPM7 mRNA is highly expressed in adipose tissue, however, its functional expression in adipocytes remains to be elucidated. In this study, we demonstrated for the first time that TRPM7 was functionally expressed in both freshly isolated white adipocytes and in 3T3-L1 adipocytes differentiated from a 3T3-L1 pre-adipocyte cell line by whole-cell patch-clamp recordings. Consistent with known properties of TRPM7 current, the current in adipocytes was activated by the elimination of extracellular divalent cations and the reduction of intracellular free Mg2+ concentrations, and was inhibited by the TRPM7 inhibitors, 2-aminoethyl diphenylborinate (2-APB), hydrogen peroxide (H2 O2 ), N-methyl maleimide (NMM), NS8593, and 2-amino-2-[2-(4-octylphenyl)ethyl]-1,3-propanediol (FTY720). Treatment with small-interfering (si) RNA targeting TRPM7 resulted in a reduction in the current to 23 ± 7% of nontargeting siRNA-treated adipocytes. Moreover a TRPM7 activator, naltriben, increased the TRPM7-like current and [Ca2+ ]i in 3T3-L1 adipocytes but not in TRPM7-knockdown adipocytes. These findings indicate that TRPM7 is functionally expressed, and plays a role as a Ca2+ influx pathway in adipocytes.
9 citations
TL;DR: The results of current study indicated that extracellular choline is primarily transported via CTL1, relying on a direct H+ gradient that functions as a driving force in Fa2N-4 cells, which provides further insights into the pathogenesis of liver disease involving choline metabolism.
Abstract: Choline is used to synthesize phospholipids and a lack of choline induces a number of liver‑related diseases, including non‑alcoholic steatohepatitis. The current study characterized the choline uptake system, at molecular and functional levels, in the immortalized human hepatic cell line, Fa2N‑4, to identify the specific choline transporter involved in choline uptake. The present study also assesed whether choline deficiency or the inhibited choline uptake affected cell viability and apoptosis. Reverse transcription‑quantitative polymerase chain reaction (PCR) revealed choline transporter‑like protein 1 (CTL1) and CTL2 mRNA and protein expression in Fa2N‑4 cells. [Methyl‑3H]choline studies revealed choline uptake was saturable and mediated by a single transport system that functioned in a Na+‑independent but pH‑dependent manner, which was similar to CTL1. Hemicholinium‑3 (HC‑3), which is a choline uptake inhibitor, and choline deficiency inhibited cell viability, increased caspase‑3 and ‑7 activities, and increased fluorescein isothiocyanate‑Annexin V immunofluorescent staining indicated apoptosis. Immunofluorescent staining also revealed CTL1 and CTL2 localized in plasma and mitochondrial membranes, respectively. [Methyl‑3H]choline uptake was enhanced by a protein kinase C (PKC) activator, phorbol‑12‑myristate 13‑acetate (PMA). Immunofluorescence staining and western blot analysis demonstrated increased CTL1 expression on the cell membrane following PMA treatment. The results of current study indicated that extracellular choline is primarily transported via CTL1, relying on a direct H+ gradient that functions as a driving force in Fa2N‑4 cells. Furthermore, it was hypothesized that CTL1 and the choline uptake system are strongly associated with cell survival, and that the choline uptake system is modulated by PKC signaling via increased CTL1 expression on the cell surface. These findings provide further insights into the pathogenesis of liver disease involving choline metabolism.
4 citations