The regulation of glycine transporter GLYT1 is mainly mediated by protein kinase Calpha in C6 glioma cells.
ABSTRACT Glycine has been shown to possess important functions as a bidirectional neurotransmitter. At synaptic clefts, the concentration of glycine is tightly regulated by the uptake of glycine released from nerve terminals into glial cells by the transporter GLYT1. It has been recently demonstrated that protein kinase C (PKC) mediates the downregulation of GLYT1 activity in several cell systems. However, it remains to be elucidated which subtypes of PKC might be important in the regulation of GLYT1 activity. In this study, we attempted to make clear the mechanism of the phorbol 12-myristate 13-acetate (PMA)-suppressed uptake of glycine in C6 glioma cells which have the native expression of GLYT1. In C6 cells, the expression of PKCalpha, PKCdelta, and PKCvarepsilon of the PMA-activated subtypes was detected. The PMA-suppressed action was fully reversed by the removal of both extracellular and intracellular Ca(2+). Furthermore, the inhibitory effects of PMA or thymeleatoxin (THX), which is a selective activator of conventional PKC (cPKC), were blocked by the downregulation of all PKCs expressed in C6 cells by long-term incubation with THX, or pretreatment with GF109203X or Gö6983, which are broad inhibitors of PKC, or Gö6976, a selective inhibitor of cPKC. On the other hand, treatment of C6 cells with ingenol, a selective activator of novel PKCs, especially PKCdelta and PKCvarepsilon, did not affect the transport of glycine. Silencing of PKCdelta expression by using RNA interference or pretreatment with the inhibitor peptide for PKCvarepsilon had no effect on the PMA-suppressed uptake of glycine. Together, these results suggest PKCalpha to be a crucial factor in the regulation of glycine transport in C6 cells.
- SourceAvailable from: Lise Sofie Haug Nissen-Meyer[Show abstract] [Hide abstract]
ABSTRACT: The system N transporter SN1 (also known as SNAT3) is enriched on perisynaptic astroglial cell membranes. SN1 mediates electroneutral and bidirectional glutamine transport, and regulates the intracellular as well as the extracellular concentrations of glutamine. We hypothesize that SN1 participates in the glutamate/γ-aminobutyric acid (GABA)-glutamine cycle and regulates the amount of glutamine supplied to the neurons for replenishment of the neurotransmitter pools of glutamate and GABA. We also hypothesize that its activity on the plasma membrane is regulated by protein kinase C (PKC)-mediated phosphorylation and that SN1 activity has an impact on synaptic plasticity. This review discusses reports on the regulation of SN1 by PKC and presents a consolidated model for regulation and degradation of SN1 and the subsequent functional implications. As SN1 function is likely also regulated by PKC-mediated phosphorylation in peripheral organs, the same mechanisms may, thus, have impact on e.g., pH regulation in the kidney, urea formation in the liver, and insulin secretion in the pancreas.Frontiers in Endocrinology 01/2013; 4:138.
- Radiotherapy and Oncology 05/2011; 99. · 4.52 Impact Factor
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ABSTRACT: Spinal astrocytes have important mechanistic contributions to the initiation and maintenance of neurodegenerative diseases and chronic pain. Under inflammatory conditions, spinal astrocytes are exposed to cytokines such as tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ), and these cytokines could alter astrocytic function by modulating connexin (Cx43), subunits that form channels that modulate intercellular communication in astrocytes. The current study investigated the alteration of Cx43-gap junction in rat primary cultured spinal astrocytes stimulated with cytokines by real-time PCR and Western blotting. The transcriptional and translational levels of Cx43 were significantly but partially reduced 24 and 48 hr treatment with either TNF-α (10 ng/ml) or IFN-γ (5 ng/ml). A mixture of TNF-α and IFN-γ led to a robust decrease of Cx43 expression and, moreover, a moderate reduction of gap junction intercellular communication (GJIC), which was evaluated by a scrap loading/dye transfer assay. Both the decrease of Cx43 expression and the reduction in GJIC induced by the mixture of TNF-α and IFN-γ were prevented by blocking c-jun terminal kinase (JNK) but not by blocking extracellular signaling molecules ERK and p38 kinase, indicating a specific role of astrocytic JNK in the response to cytokines. In addition, treatment with cytokines potently induced the phosphorylation of JNK and c-jun in a time-dependent manner. These results indicate that intercellular communication of astrocytes is significantly disrupted in the inflammatory state and that stimulation of spinal astrocytes with inflammatory cytokines leads to significant inhibition of Cx43-GJIC through activation of the JNK signaling pathway. © 2013 Wiley Periodicals, Inc.Journal of Neuroscience Research 03/2013; · 2.97 Impact Factor