Conference Publication Details
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KAAR, A., LEAHY, J., O’DONOGHUE, K. & RAE, M.G
Neuroscience Ireland
Evidence for the involvement of ryanodine receptors in Ca2+ release by group I metabotropic glutamate receptors in cultured rat hippocampal neurons
2013
September
Validated
1
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Group 1 metabotropic glutamate receptors (I-mGluRs) are important neuromodulators of synaptic plasticity and also play a role in the aetiology of Alzheimer’s disease. For many years I-mGluRs have been thought to mediate their effects primarily by mobilising intracellular calcium release from the neuronal endoplasmic reticulum (ER) via the production of inositol-1,4,5-trisphosphate (IP3), catalysed by the activation of phospholipase C beta (PLC). However, recent work suggests that in acutely dissociated rat hippocampal neurons I- mGluR-mediated intracellular Ca2+ mobilisation is at least partly dependent on cyclic ADP ribose (cADPR)-dependent activation of ryanodine receptors (RyRs)1. It was the aim therefore of the current study to determine if I-mGluR–mediated Ca2+ signals within cultured rat hippocampal neurons, previously ascribed to activation of the PLC/IP3 signalling pathway2, may be mediated via the aforementioned cADPR/RyR pathway. Experiments were carried out utilising cultured hippocampal neurons obtained from 3-5 day old Sprague-Dawley rat pups as described previously2. Conventional calcium imaging was used to record intracellular somatic calcium elevations in neurons loaded with the calcium-sensitive dye, fluo-2AM (150 microM). Experiments were carried out at room temperature with neurons continuously perfused (2ml/min) with a standard HEPES-buffered saline solution (HBSS). All drugs were added to the perfusate. Data are expressed as means ± S.E.M. Somatic I-mGluR–mediated [Ca2+]i signals, evoked using the specific agonist, (S)-3,5 dihydroxyphenylglycine (DHPG; 50 microM; 2 min), were significantly enhanced (as determined by measuring area under the curve) relative to control following depolarisation with elevated extracellular K+ (15 mM)-containing HBSS solution, by 26.1±9.5% (P<0.05; n=60) in line with previous studies on rat hippocampal neurones2. The cADPR antagonist, nicotinamide (5 mM), and the RyR antagonist dantrolene (10 microM) each significantly reduced the area of the DHPG-evoked [Ca2+]i signals elicited in 15 mM K+-HBSS by 24±8% (n=34; P<0.01) and 18±4% (n=61; P<0.01), respectively. Application of the PLC inhibitor, U73122 (5 microM) also significantly inhibited the DHPG-evoked [Ca2+]i signals by 56.5±16.3% (n=9; P<0.05). Co-application of U73122 and nicotinamide completely abolished DHPG-evoked [Ca2+]i signals (n=10; P<0.05). These results support the suggestion that I-mGluR-evoked neuronal [Ca2+]i signals may be mediated, at least in part, via cADPR-dependent RyR signalling. References 1. Sohn JW et al. (2011) Cyclic ADP ribose-dependent Ca2+ release by group I metabotropic glutamate receptors in acutely dissociated rat hippocampal neurons. PloS one 6, e26625. 2. Rae MG et al. (2000) Role of Ca2+ stores in metabotropic L-glutamate receptor-mediated supralinear Ca2+ signaling in rat hippocampal neurons. J. Neurosci. 20, 8628-8636.
Grant Details
The Physiological Society