Elsevier

Experimental Neurology

Volume 197, Issue 2, February 2006, Pages 495-504
Experimental Neurology

Regular Article
NMDA receptor-mediated epileptiform persistent activity requires calcium release from intracellular stores in prefrontal neurons

https://doi.org/10.1016/j.expneurol.2005.05.018Get rights and content

Abstract

Various normal and pathological forms of synchronized population activity are generated by recurrent excitation among pyramidal neurons in the neocortex. However, the intracellular signaling mechanisms underlying this activity remain poorly understood. In this study, we have examined the cellular properties of synchronized epileptiform activity in the prefrontal cortex with particular emphasis on a potential role of intracellular calcium stores. We find that the zero-magnesium-induced synchronized activity is blocked by inhibition of sarco-endoplasmic reticulum Ca2+-ATPases, phospholipase C (PLC), the inositol 1,4,5-trisphosphate (IP3) receptor, and the ryanodine receptor. This same activity is, however, not affected by application of metabotropic glutamatergic receptor (mGluR) agonists, nor by introduction of an mGluR antagonist. These results suggest that persistent synchronized activity in vitro is dependent upon calcium release from internal calcium stores through the activation of PLC-IP3 receptor pathway. Our findings also raise the possibility that intracellular calcium release may be involved in the generation of pathologic synchronized activity in epilepsy in vivo and in physiological forms of synchronized cortical activity.

Introduction

Neural assemblies in a variety of brain areas display synchronized oscillations in response to sensory stimuli, and this synchronous activity has been suggested to underlie cognitive functions (Singer, 1993, Salinas and Sejnowski, 2001, Varela et al., 2001). An essential basis of such oscillation in the neocortex is thought to be action potential-dependent excitation among interconnected pyramidal neurons participating in local microcircuits (Steriade et al., 1993, Gray and McCormick, 1996, Sanchez-Vives and McCormick, 2000, Salinas and Sejnowski, 2001). In the prefrontal cortex (PFC) in particular, recurrent excitatory interactions are thought to account for synchronized persistent activity underlying the neural process of working memory (Goldman-Rakic, 1995).

Recurrent excitation in cortical networks is mediated by glutamatergic excitation and balanced by gamma aminobutyric acid (GABA)-ergic inhibition. Disinhibition of in vitro cortical slice preparations by blockade of GABAA receptors or enhancement of N-methyl-d-aspartic acid (NMDA)-mediated recurrent excitation with the removal of magnesium (Mg2+) from the extracellular medium causes spontaneous synchronous burst firing which has been extensively used as models of epileptic activity (Connors, 1984, Chagnac-Amitai and Connors, 1989, Avoli et al., 1991, Traub et al., 1994, Telfeian and Connors, 1999, McCormick and Contreras, 2001, Avoli et al., 2002, Castro-Alamancos and Rigas, 2002, Cohen et al., 2002, Stoop et al., 2003).

Several lines of evidence indicate that release of intracellular Ca2+ stores is related to synchronous activity in cortical pyramidal neurons. For example, stimulation of the NMDA receptor to generate epileptiform activity has been shown to mobilize internal Ca2+ stores (Yoshimura et al., 2001). Moreover, activation of group I mGluRs through high frequency stimulation of excitatory afferents elicits propagating Ca2+ waves that are dependent on IP3Rs (Nakamura et al., 2002). Although the physiological effects of the Ca2+ waves on the excitability of cortical neurons is not clear, it is likely that the high concentration of Ca2+ ions associated with these waves regulates the activity of numerous Ca2+-dependent ion channels underlying inward and/or outward currents. Conversely, the population activity of pyramidal neurons during epileptiform bursting is very similar to that previously determined to be sufficient to elicit regenerative intracellular Ca2+ release (Nakamura et al., 2000, Larkum et al., 2003). Thus, population activity may also contribute to intracellular Ca2+ release and, as a result, perpetuate burst firing. How does intracellular Ca2+ release regulate synchronized population activity? We hypothesize that a large amount of Ca2+ could enter the cell through the activated NMDARs in the neuronal network, which in turn contribute to the production of IP3 by activating Ca2+-sensitive PLC isoforms, resulting in activation and opening of IP3Rs and/or RyRs in the ER to release sequestered Ca2+ into the cytosol (Berridge et al., 2000). Here, we tested this hypothesis and examined the effects of pharmacologic agents that inhibit release of intracellular Ca2+ stores on the physiological properties of synchronized activity as it strengthens and evolves into persistent epileptiform activity in nominally Mg2+-free solution in slice preparation of the PFC. We found that Ca2+ release from internal stores was essential for generation of persistent epileptiform activity in layer V pyramidal cells of the PFC.

Section snippets

Slices preparation and physiological recording

Experiments were performed on layer V pyramidal neurons from medial prefrontal cortical slices prepared from 21 ferrets at ages of 7–14 weeks. Animals were treated according to guidelines of National Institutes of Health and in accordance with Yale’s Institutional Animal Care and Use Committee regulations. The animals were deeply anesthetized with sodium pentobarbital (100 mg/kg), and the brains were removed and immediately placed in ice-cold oxygenated artificial cerebrospinal fluid (ACSF)

Results

Multiple whole-cell recordings were performed in a total of 92 layer V pyramidal neurons in the medial PFC. Pyramidal cells were visualized and identified with the assistance of IR-DIC and on the basis of regular or intrinsic bursting firing patterns. The morphologies of recorded neurons were confirmed by biocytin labeling (data not shown). Zero-Mg2+ extracellular solution was applied to relieve the blockade of NMDA receptor channels and to induce synchronized bursting that transformed into

Discussion

We have employed multiple whole-cell recording combined with pharmacological applications to address the intracellular signaling mechanisms of synchronized activity induced by transient removal of extracellular Mg2+ in the prefrontal cortex. Application of glutamate receptor agonists and antagonists indicated that the synchronized population activities were generated by recurrent synaptic activity mediated by NMDARs and AMPARs, and were not modulated by mGluRs. Pharmacological manipulation of

Acknowledgments

This paper is dedicated to the memory of Dr. Patricia S. Goldman-Rakic, a great neuroscientist and mentor. We are grateful to Drs. L.D. Selemon, T. Koos, C.D. Paspalas, and N. Kabbani for comments on the manuscript. This work was supported by P50 MH44866, RO1 MH38546, Essel Foundation to PGR, and NARSAD Young Investigator Award to WJG.

References (63)

  • S. Pal et al.

    Epileptogenesis induces long-term alterations in intracellular calcium release and sequestration mechanisms in the hippocampal neuronal culture model of epilepsy

    Cell Calcium

    (2001)
  • N.W. Seidler et al.

    Cyclopiazonic acid is a specific inhibitor of the Ca2+-ATPase of sarcoplasmic reticulum

    J. Biol. Chem.

    (1989)
  • M. Treiman et al.

    A tool coming of age: thapsigargin as an inhibitor of sarco-endoplasmic reticulum Ca(2+)-ATPases

    Trends Pharmacol. Sci.

    (1998)
  • X. Wang et al.

    Mechanism of synchronized Ca2+ oscillations in cortical neurons

    Brain Res.

    (1997)
  • H. Yoshimura et al.

    Synchronized population oscillation of excitatory synaptic potentials dependent of calcium-induced calcium release in rat neocortex layer II/III neurons

    Brain Res.

    (2001)
  • M. Avoli et al.

    Epileptiform activity induced by low extracellular magnesium in the human cortex maintained in vitro

    Ann. Neurol.

    (1991)
  • T. Badea et al.

    Calcium imaging of epileptiform events with single-cell resolution

    J. Neurobiol.

    (2001)
  • S. Bardo et al.

    Presynaptic internal Ca2+ stores contribute to inhibitory neurotransmitter release onto mouse cerebellar Purkinje cells

    Br. J. Pharmacol.

    (2002)
  • M.J. Berridge et al.

    The versatility and universality of calcium signalling

    Nat. Rev., Mol. Cell Biol.

    (2000)
  • O. Caillard et al.

    Activation of presynaptic and postsynaptic ryanodine-sensitive calcium stores is required for the induction of long-term depression at GABAergic synapses in the neonatal rat hippocampus amphetamine

    J. Neurosci.

    (2000)
  • A.G. Carter et al.

    Assessing the role of calcium-induced calcium release in short-term presynaptic plasticity at excitatory central synapses

    J. Neurosci.

    (2002)
  • M.A. Castro-Alamancos et al.

    Synchronized oscillations caused by disinhibition in rodent neocortex are generated by recurrent synaptic activity mediated by AMPA receptors

    J. Physiol.

    (2002)
  • Y. Chagnac-Amitai et al.

    Synchronized excitation and inhibition driven by intrinsically bursting neurons in neocortex

    J. Neurophysiol.

    (1989)
  • I. Cohen et al.

    On the origin of interictal activity in human temporal lobe epilepsy in vitro

    Science

    (2002)
  • B.W. Connors

    Initiation of synchronized neuronal bursting in neocortex

    Nature

    (1984)
  • R. Cossart et al.

    Attractor dynamics of network UP states in the neocortex

    Nature

    (2003)
  • P. De Smet et al.

    Xestospongin C is an equally potent inhibitor of the inositol 1,4,5-trisphosphate receptor and the endoplasmic-reticulum Ca(2+) pumps

    Cell Calcium

    (1999)
  • B.E. Ehrlich et al.

    Intracellular calcium release channels

    Chin. J. Physiol.

    (1994)
  • M. Fill et al.

    Ryanodine receptor calcium release channels

    Physiol. Rev.

    (2002)
  • M. Galante et al.

    Presynaptic ryanodine-sensitive calcium stores contribute to evoked neurotransmitter release at the basket cell-Purkinje cell synapse

    J. Neurosci.

    (2003)
  • W.J. Gao et al.

    Selective modulation of excitatory and inhibitory microcircuits by dopamine

    Proc. Natl. Acad. Sci. U. S. A.

    (2003)
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