Altered expression of connexin subtypes in mesial temporal lobe epilepsy in humans.

OBJECT The causes of epileptic events remain unclear. Much in vitro and in vivo experimental evidence suggests that gap junctions formed by connexins (Cxs) between neurons and/or astrocytes contribute to the generation and maintenance of seizures; however, few experiments have been conducted in humans, and those completed have shown controversial data. The authors designed a study to compare the level of expression of Cxs in hippocampi from epileptic and nonepileptic patients to assess whether an alteration of gap junction expression in epileptic tissue plays a role in seizure origin and propagation. METHODS The expression of Cxs32, -36, and -43 was studied in 47 consecutive samples of hippocampi obtained from epileptic patients who had undergone an amygdalohippocampectomy for the treatment of intractable seizure. These expression levels were compared with those in hippocampi obtained in nonepileptic patients during postmortem dissection. Immunostaining was performed to create one slide for each of the three Cxs. Each slide demonstrated multiple cells from each of six regions (CA1, CA2, CA3, CA4, dentate gyrus, and subiculum). Two independent reviewers rated each Cx-region combination according to an immunoreactive score. Across all three measures-that is, staining intensity, percentage of positively stained cells, and immunoreactive score-Cx32 appeared to be expressed at a significantly lower level in the epileptic patients compared with controls (p < 0.001 for each measure), whereas Cx43 appeared to be expressed more among the epileptic patients (p < 0.001 for each measure). There was no evidence of any differential expression of Cx36. There was, however, regional variation within each Cx subtype. For Cx36, the staining intensity was higher in the CA2 region in the epilepsy group. CONCLUSIONS The increase in Cx43, decrease in Cx32, and preservation of Cx36 expression in hippocampi from epileptic persons could play a role in the development of seizures in patients with temporal sclerosis. Additional studies must be completed to understand this mechanism better.

[1]  P. Micevych,et al.  Distribution of mRNAs coding for liver and heart gap junction proteins in the rat central nervous system , 1991, The Journal of comparative neurology.

[2]  Andreas Draguhn,et al.  Reduction of high‐frequency network oscillations (ripples) and pathological network discharges in hippocampal slices from connexin 36‐deficient mice , 2002, The Journal of physiology.

[3]  J. Bechberger,et al.  Regional differences in connexin32 and connexin43 messenger RNAs in rat brain , 1990, Neuroscience Letters.

[4]  C. Bruton,et al.  The neuropathology of temporal lobe epilepsy , 1988 .

[5]  CR Houser,et al.  Altered patterns of dynorphin immunoreactivity suggest mossy fiber reorganization in human hippocampal epilepsy , 1990, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[6]  The balance between excitation and inhibition in dentate granule cells and its role in epilepsy. , 1992, Epilepsy research. Supplement.

[7]  Helen J. Cross,et al.  A Possible Role for Gap Junctions in Generation of Very Fast EEG Oscillations Preceding the Onset of, and Perhaps Initiating, Seizures , 2001, Epilepsia.

[8]  F. Dudek,et al.  Role of electrical interactions in synchronization of epileptiform bursts. , 1986, Advances in neurology.

[9]  Carissa G. Fonseca,et al.  Upregulation in astrocytic connexin 43 gap junction levels may exacerbate generalized seizures in mesial temporal lobe epilepsy , 2002, Brain Research.

[10]  K. Willecke,et al.  Differential expression of three gap junction proteins in developing and mature brain tissues. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[11]  R. Traub,et al.  Axonal Gap Junctions Between Principal Neurons: A Novel Source of Network Oscillations, and Perhaps Epileptogenesis , 2002, Reviews in the neurosciences.

[12]  G. Tamás,et al.  Cholinergic activation and tonic excitation induce persistent gamma oscillations in mouse somatosensory cortex in vitro , 1998, The Journal of physiology.

[13]  J. Degen,et al.  The murine gap junction gene connexin36 is highly expressed in mouse retina and regulated during brain development , 1998, FEBS letters.

[14]  A C Campos de Carvalho,et al.  Gap-junctional coupling between neurons and astrocytes in primary central nervous system cultures. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[15]  F. Dudek,et al.  Prolonged field bursts in the dentate gyrus: dependence on low calcium, high potassium, and nonsynaptic mechanisms. , 1992, Journal of neurophysiology.

[16]  J. C. Nelson,et al.  Neurotransmitter Coupling through Gap Junctions in the Retina , 1998, The Journal of Neuroscience.

[17]  D. Tauck,et al.  Evidence of functional mossy fiber sprouting in hippocampal formation of kainic acid-treated rats , 1985, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[18]  D. Spray,et al.  Gap junctions in glia. Types, roles, and plasticity. , 1999, Advances in experimental medicine and biology.

[19]  Corsellis Ja,et al.  Neuropathology of status epilepticus in humans. , 1983, Advances in neurology.

[20]  E. Hertzberg,et al.  LM and EM immunolocalization of the gap junctional protein connexin 43 in rat brain , 1990, Brain Research.

[21]  P. Carlen,et al.  Upregulation of gap junction connexin 32 with epileptiform activity in the isolated mouse hippocampus , 2001, Neuroscience.

[22]  R. Traub,et al.  Virus‐induced electrotonic coupling: Hypothesis on the mechanism of periodic EEG discharges in Creutzfeldt‐Jakob disease , 1981, Annals of neurology.

[23]  Moshe Kushnir,et al.  The role of gap junctions in seizures , 2000, Brain Research Reviews.

[24]  E. Kandel,et al.  Epileptogenic agents enhance transmission at an identified weak electrical synapse in Aplysia. , 1981, Science.

[25]  D. Goodenough,et al.  Diverse functions of vertebrate gap junctions. , 1998, Trends in cell biology.

[26]  R. Traub,et al.  Gap junctions, fast oscillations and the initiation of seizures. , 2004, Advances in experimental medicine and biology.

[27]  T. Powell,et al.  Gap junctions between dendrites and somata of neurons in the primate sensori-motor cortex , 1978, Proceedings of the Royal Society of London. Series B. Biological Sciences.

[28]  G. Ivy,et al.  Astrocytes in kindling: relevance to epileptogenesis , 1996, Epilepsy Research.

[29]  D. Spencer,et al.  Astrocytes from Human Hippocampal Epileptogenic Foci Exhibit Action Potential–Like Responses , 1998, Epilepsia.

[30]  E. Mugnaini,et al.  Cell junctions and intramembrane particles of astrocytes and oligodendrocytes: A freeze-fracture study , 1982, Neuroscience.

[31]  D. Spray,et al.  Gap junctions in the brain: where, what type, how many and why? , 1993, Trends in Neurosciences.

[32]  R. S. Sloviter,et al.  The functional organization of the hippocampal dentate gyrus and its relevance to the pathogenesis of temporal lobe epilepsy , 1994, Annals of neurology.

[33]  S. B. Kater,et al.  Evidence for glutamate-mediated activation of hippocampal neurons by glial calcium waves. , 1995, Journal of neurobiology.

[34]  R. Traub,et al.  Neuronal networks for induced ‘40 Hz’ rhythms , 1996, Trends in Neurosciences.

[35]  Kenneth D Harris,et al.  Selective Impairment of Hippocampal Gamma Oscillations in Connexin-36 Knock-Out Mouse In Vivo , 2003, The Journal of Neuroscience.

[36]  F. Dudek,et al.  Electrophysiology of GABA-mediated synaptic transmission and possible roles in epilepsy , 1991, Neurochemical Research.

[37]  A. Cornell-Bell,et al.  Human epileptic astrocytes exhibit increased gap junction coupling , 1995, Glia.

[38]  J. Cavazos,et al.  Long‐term structural and functional alterations induced in the hippocampus by kindling: Implications for memory dysfunction and the development of epilepsy , 1994, Hippocampus.

[39]  E. Hertzberg,et al.  Gap junction protein in rat hippocampus: Correlative light and electron microscope immunohistochemical localization , 1989, The Journal of comparative neurology.

[40]  F. Dudek,et al.  ‘NON‐SYNAPTIC’ MECHANISMS IN SEIZURES AND EPILEPTOGENESIS , 1998, Cell biology international.

[41]  A. Konnerth,et al.  Slow transmission of neural activity in hippocampal area CA1 in absence of active chemical synapses , 1984, Nature.

[42]  O. Paulsen,et al.  Cholinergic induction of network oscillations at 40 Hz in the hippocampus in vitro , 1998, Nature.

[43]  N. Belluardo,et al.  Cloning of a new gap junction gene (Cx36) highly expressed in mammalian brain neurons , 1998, The European journal of neuroscience.

[44]  Miles A. Whittington,et al.  Impaired Electrical Signaling Disrupts Gamma Frequency Oscillations in Connexin 36-Deficient Mice , 2001, Neuron.

[45]  D. Feldmeyer,et al.  Connexin expression in electrically coupled postnatal rat brain neurons. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[46]  J. Jefferys,et al.  Nonsynaptic modulation of neuronal activity in the brain: electric currents and extracellular ions. , 1995, Physiological reviews.

[47]  F. Dudek,et al.  A physiological test for electrotonic coupling between CA1 pyramidal cells in rat hippocampal slices , 1982, Brain Research.

[48]  R. Traub,et al.  Synchronized oscillations in interneuron networks driven by metabotropic glutamate receptor activation , 1995, Nature.

[49]  M. P. Witter,et al.  Connectivity of the rat hippocampus , 1989 .

[50]  L. Hertz,et al.  Functional interactions between neurons and astrocytes. II. Potassium homeostasis at the cellular level , 1983, Progress in Neurobiology.

[51]  D. Paul,et al.  Genetic diseases and gene knockouts reveal diverse connexin functions. , 1999, Annual review of physiology.

[52]  F. Dudek,et al.  Synchronous neural afterdischarges in rat hippocampal slices without active chemical synapses. , 1982, Science.

[53]  R. Jabs,et al.  Functional and Molecular Properties of Human Astrocytes in Acute Hippocampal Slices Obtained from Patients with Temporal Lobe Epilepsy , 2000, Epilepsia.

[54]  M. Ishimatsu,et al.  Synchronous Activity in Locus Coeruleus Results from Dendritic Interactions in Pericoerulear Regions , 1996, The Journal of Neuroscience.

[55]  F. L. Glo¨tzner.  Membrane properties of neuroglia in epileptogenic gliosis , 1973 .

[56]  J. Parnavelas,et al.  Gap junctions in the adult cerebral cortex: Regional differences in their distribution and cellular expression of connexins , 1996, The Journal of comparative neurology.

[57]  D. Paul.  Molecular cloning of cDNA for rat liver gap junction protein , 1986, The Journal of cell biology.

[58]  D. Spray.  MOLECULAR PHYSIOLOGY OF GAP JUNCTION CHANNELS , 1996, Clinical and experimental pharmacology & physiology.

[59]  Y. Isomura,et al.  Excitatory gaba input directly drives seizure-like rhythmic synchronization in mature hippocampal CA1 pyramidal cells , 2003, Neuroscience.

[60]  R. Bruzzone,et al.  Connections with connexins: the molecular basis of direct intercellular signaling. , 1996, European journal of biochemistry.

[61]  D. Spray,et al.  Gap Junctions in Glia , 1999 .

[62]  F. Dudek,et al.  Mechanisms of neuronal synchronization during epileptiform activity. , 1999, Advances in neurology.

[63]  F. Dudek,et al.  Dye-coupling between CA3 pyramidal cells in slices of rat hippocampus , 1980, Brain Research.

[64]  R. Traub,et al.  Electrical coupling underlies high-frequency oscillations in the hippocampus in vitro , 1998, Nature.

[65]  H. Beck,et al.  Expression of connexin genes in hippocampus of kainate-treated and kindled rats under conditions of experimental epilepsy. , 2000, Brain research. Molecular brain research.

[66]  E. Hertzberg,et al.  Gap junctions: New tools, new answers, new questions , 1991, Neuron.

[67]  J. Jefferys,et al.  Low‐calcium field burst discharges of CA1 pyramidal neurones in rat hippocampal slices. , 1984, The Journal of physiology.

[68]  David L. Paul,et al.  Gap junction gene expression in human seizure disorder , 1991, Experimental Neurology.

[69]  R. Llinás,et al.  Electrotonic coupling between neurons in cat inferior olive. , 1974, Journal of neurophysiology.

[70]  E J Vigmond,et al.  Mechanisms of electrical coupling between pyramidal cells. , 1997, Journal of neurophysiology.

[71]  D. Gros,et al.  Immunological characterization of rat cardiac gap junctions: Presence of common antigenic determinants in heart of other vertebrate species and in various organs , 1988, The Journal of Membrane Biology.

[72]  M. Gutnick,et al.  Dye coupling and possible electrotonic coupling in the guinea pig neocortical slice. , 1981, Science.