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Protein phosphatase modulation of somatostatin receptor signaling in the mouse hippocampus.

ABSTRACT: Many inhibitory interneurones in the hippocampus release the neuropeptide somatostatin (SST) which inhibits neuronal excitability through Gi/Go-coupled receptors. To investigate the signaling pathways underlying the SST inhibition of neuronal excitability in the hippocampus, we performed perforated patch-clamp recordings from CA1 pyramidal neurones in acute brain slices from P14-P18 mice. Bath application of 1 ?M SST reversibly reduces the frequency of action potential firing in response to depolarising current steps, and is associated with neuronal hyperpolarisation and a reduction in membrane resistance. This effect is mediated by potassium channels with KCNK-like pharmacology. In addition, in slices that have been cultured in vitro for seven days or more, SST also produces a hyperpolarisation independent reduction in action potential firing, which can be also observed in acute slices when the Ser/Thr protein phosphatases PP2A and PP4 are inhibited selectively with fostriecin. This hyperpolarisation independent effect of SST appears to be mediated by G-protein-activated inwardly rectifying K+ (GIRK) channels. Knockdown of protein phosphatase 5, by Cre recombinase mediated deletion of the floxed Ppp5c gene, blocks the hyperpolarisation independent effect of SST, and reduces the hyperpolarisation dependent effect in a manner consistent with increased SST receptor desensitisation. Thus, reversible protein phosphorylation provides a mechanism to enhance or diminish the inhibitory effect of SST, which could allow system level regulation of circuit excitability in the hippocampus.

SUBMITTER: Lucas SJ

PROVIDER: S-EPMC4655177 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

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Protein phosphatase modulation of somatostatin receptor signaling in the mouse hippocampus.

Lucas Sarah J SJ Armstrong David L DL

Neuropharmacology 20150718

Many inhibitory interneurones in the hippocampus release the neuropeptide somatostatin (SST) which inhibits neuronal excitability through Gi/Go-coupled receptors. To investigate the signaling pathways underlying the SST inhibition of neuronal excitability in the hippocampus, we performed perforated patch-clamp recordings from CA1 pyramidal neurones in acute brain slices from P14-P18 mice. Bath application of 1 μM SST reversibly reduces the frequency of action potential firing in response to depo ...[more]

PMID: 26196943

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Project description:Fms-like tyrosine kinase 3 (FLT3) plays an important role in hematopoietic differentiation, and constitutively active FLT3 mutant proteins contribute to the development of acute myeloid leukemia. Little is known about the protein-tyrosine phosphatases (PTP) affecting the signaling activity of FLT3. To identify such PTP, myeloid cells expressing wild type FLT3 were infected with a panel of lentiviral pseudotypes carrying shRNA expression cassettes targeting different PTP. Out of 20 PTP tested, expressed in hematopoietic cells, or presumed to be involved in oncogenesis or tumor suppression, DEP-1 (PTPRJ) was identified as a PTP negatively regulating FLT3 phosphorylation and signaling. Stable 32D myeloid cell lines with strongly reduced DEP-1 levels showed site-selective hyperphosphorylation of FLT3. In particular, the sites pTyr-589, pTyr-591, and pTyr-842 involved in the FLT3 ligand (FL)-mediated activation of FLT3 were hyperphosphorylated the most. Similarly, acute depletion of DEP-1 in the human AML cell line THP-1 caused elevated FLT3 phosphorylation. Direct interaction of DEP-1 and FLT3 was demonstrated by "substrate trapping" experiments showing association of DEP-1 D1205A or C1239S mutant proteins with FLT3 by co-immunoprecipitation. Moreover, activated FLT3 could be dephosphorylated by recombinant DEP-1 in vitro. Enhanced FLT3 phosphorylation in DEP-1-depleted cells was accompanied by enhanced FLT3-dependent activation of ERK and cell proliferation. Stable overexpression of DEP-1 in 32D cells and transient overexpression with FLT3 in HEK293 cells resulted in reduction of FL-mediated FLT3 signaling activity. Furthermore, FL-stimulated colony formation of 32D cells expressing FLT3 in methylcellulose was induced in response to shRNA-mediated DEP-1 knockdown. This transforming effect of DEP-1 knockdown was consistent with a moderately increased activation of STAT5 upon FL stimulation but did not translate into myeloproliferative disease formation in the 32D-C3H/HeJ mouse model. The data indicate that DEP-1 is negatively regulating FLT3 signaling activity and that its loss may contribute to but is not sufficient for leukemogenic cell transformation.

Dataset Information

Protein phosphatase modulation of somatostatin receptor signaling in the mouse hippocampus.

Publications

Protein phosphatase modulation of somatostatin receptor signaling in the mouse hippocampus.

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