Lability of GABAA receptor function in human partial epilepsy: possible relationship to hypometabolism.

The function of the gamma-aminobutyric acid type A receptor (GABA(A)R) is maintained by endogenous phosphorylation. We have shown that the corresponding kinase is the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH), using the locally produced glycolytic ATP. In addition, using cerebral tissue obtained during curative surgery for epilepsy, we showed that both the endogenous phosphorylation and the GABA(A)R function are significantly reduced in the "epileptogenic" cerebral cortex when compared to "control" tissue. This dysfunction likely contributes to seizure generation and/or transition from the interictal to the ictal state. Glucose utilization is decreased in the epileptogenic cortex of patients with partial epilepsy in the interictal state, but the relationship to the disorder remains unclear. We propose that this hypometabolism is related to the deficiency in the endogenous phosphorylation of GABA(A)R and the resulting greater lability of GABAergic inhibition. Several lines of evidences indeed suggest that GABAergic inhibition is costly in terms of metabolic consumption. The deficiency of this glycolysis-dependent mechanism may thus link epileptogenicity to glucose hypometabolism. The antiepileptic effect of ketogenic diets may be mediated by the subsequent rise in the NADH/NAD(+) index, which favors GABA(A)R endogenous phosphorylation and should contribute to restoration of GABAergic inhibition in the epileptogenic zone.

Histaminergic ligands improve vestibular compensation in the cat: behavioural, neurochemical and molecular evidence.

This study analysed the effects of betahistine and thioperamide, two histamine H(3) receptor antagonists, on the recovery process after unilateral vestibular neurectomy (UVN) in the cat. In UVN animals untreated or treated with betahistine or thioperamide, recovery was evaluated by recording the horizontal spontaneous nystagmus and the postural and locomotor performances. The neurochemical effects of these drugs were determined by examining their impact on the histaminergic system. We quantified the mRNA coding for histidine decarboxylase (enzyme synthesizing histamine) by in situ hybridisation in the tuberomammillary nuclei, while binding density to histamine H(3) receptors was assessed using a histamine H(3) receptor agonist ([(3)H]N-alpha-methylhistamine) and autoradiography methods in the tuberomammillary and the vestibular nuclei. Relative to the UVN-untreated group, cats treated with betahistine or thioperamide showed strongly accelerated behavioural recovery. UVN-induced 1) an up-regulation of histidine decarboxylase mRNA in the tuberomammillary nuclei, strongly accentuated under betahistine and thioperamide, 2) a reduction of the binding to histamine H(3) receptors in the vestibular and tuberomammillary nuclei, also strongly enhanced in both groups of treated cats. This study demonstrates that betahistine and thioperamide strongly improve the recovery of vestibular functions in UVN cats by interacting with the histaminergic system.

Dysfunction of GABAA receptor glycolysis-dependent modulation in human partial epilepsy.

A reduction in GABAergic neurotransmission has been put forward as a pathophysiological mechanism for human epilepsy. However, in slices of human epileptogenic neocortex, GABAergic inhibition can be clearly demonstrated. In this article we present data showing an increase in the functional lability of GABAergic inhibition in epileptogenic tissue compared with nonepileptogenic human tissue. We have previously shown that the glycolytic enzyme GAPDH is the kinase involved in the glycolysis-dependent endogenous phosphorylation of the alpha1-subunit of GABA(A) receptor, a mechanism necessary for maintaining GABA(A) function. In human epileptogenic cortex obtained during curative surgery of patients with partial seizures, we demonstrate an intrinsic deficiency of GABA(A) receptor endogenous phosphorylation resulting in an increased lability of GABAergic currents in neurons isolated from this tissue when compared with neurons from nonepileptogenic human tissue. This feature was not related to a reduction in the number of GABA(A) receptor alpha1-subunits in the epileptogenic tissue as measured by [(3)H]flunitrazepam photoaffinity labeling. Maintaining the receptor in a phosphorylated state either by favoring the endogenous phosphorylation or by inhibiting a membrane-associated phosphatase is needed to sustain GABA(A) receptor responses in epileptogenic cortex. The increased functional lability induced by the deficiency in phosphorylation can account for transient GABAergic disinhibition favoring seizure initiation and propagation. These findings imply new therapeutic approaches and suggest a functional link to the regional cerebral glucose hypometabolism observed in patients with partial epilepsy, because the dysfunctional GABAergic mechanism depends on the locally produced glycolytic ATP.

The Kell and XK proteins of the Kell blood group are not co-expressed in the central nervous system.

The Kell blood group is constituted by two covalently linked antigens at the surface of red blood cells, Kell and Kx. Whereas Kell is a metalloprotease with demonstrated in vitro enzymatic activity, the role of Kx thereon, and/or alone, remains unknown, although its absence is linked to the McLeod syndrome, a neuroacanthocytosis. In the central nervous system, the expression of Kell and XK has been suggested, but their expression patterns remain uncharacterized, as are the post-translational pathogenic mechanisms involved in the development of the McLeod syndrome. The distributions of Kell and XK were thus studied by in situ hybridization as well as immunohistochemistry in rodent and human brain. The results reveal an independent localization of the two constituents of the Kell blood group, XK (Kx) being expressed throughout this tissue, whereas Kell expression is restricted to red blood cells in cerebral vessels. The XK protein is shown to be neuronal, located mainly in intracellular compartments, suggesting a cell specific trafficking pattern, possibly associated with specific physiological functions.

Changes in the histaminergic system during vestibular compensation in the cat.

To determine how the histaminergic system is implicated in vestibular compensation, we studied the changes in histidine decarboxylase (HDC; the enzyme synthesizing histamine) mRNA regulation in the tuberomammillary (TM) nuclei of cats killed 1 week, 3 weeks and 3 months after unilateral vestibular neurectomy (UVN). We also used one- and two-step bilateral vestibular neurectomized (BVN) cats to determine whether HDC mRNA regulation depended on the asymmetrical vestibular input received by the TM nuclei neurons. In addition, we analysed the HDC mRNA changes in the TM nuclei and the recovery of behavioural functions in UVN cats treated with thioperamide, a pure histaminergic drug. Finally, we quantified binding to histamine H3 receptors (H3Rs) in the medial vestibular nucleus (VN) by means of a histamine H3R agonist ([3H]N-alpha-methylhistamine) in order to further investigate the sites and mechanisms of action of histamine in this structure. This study shows that UVN increases HDC mRNA expression in the ipsilateral TM nucleus at 1 week. This increased expression persisted 3 weeks after UVN, and regained control values at 3 months. HDC mRNA expression was unchanged in the one-step BVN cats but showed mirror asymmetrical increases in the two-step BVN compared to the 1 week UVN cats. Three weeks' thioperamide treatment induced a bilateral HDC mRNA up-regulation in the UVN cats, which was higher than in the untreated UVN group. Binding to histamine H3Rs in the MVN showed a strong bilateral decrease after thioperamide treatment, while it was reduced ipsilaterally in the UVN cats. That such changes of the histaminergic system induced by vestibular lesion and treatment may play a functional role in vestibular compensation is strongly supported by the behavioural data. Indeed, spontaneous nystagmus, posture and locomotor balance were rapidly recovered in the UVN cats treated with thioperamide. These results demonstrate that changes in histamine levels are related to vestibular compensation.

Dose- and duration-dependent effects of betahistine dihydrochloride treatment on histamine turnover in the cat.

Drugs interacting with the histaminergic system are currently used for vertigo treatment and it was shown in animal models that structural analogues of histamine like betahistine improved the recovery process after vestibular lesion. This study was aimed at determining the possible dose and duration effects of betahistine treatment on histamine turnover in normal adult cats, as judged by the level of messenger RNA for histidine decarboxylase (enzyme synthesizing histamine) in the tuberomammillary nuclei. Experiments were conducted on betahistine-treated cats receiving daily doses of 2, 5, 10, or 50 mg/kg during 1 week, 3 weeks, 2 months, or 3 months. The 1-week, 3-week, and 2- and 3-month treatments correspond to the acute, compensatory, and sustained compensatory stages of vestibular compensation, respectively. The lowest dose (2 mg/kg) given the longest time (3 months) was close to the dosage for vestibular defective patients. Data from the experimental groups were compared to control, untreated cats and to placebo-treated animals. The results clearly show that betahistine dihydrochloride administered orally in the normal cat interferes with histamine turnover by increasing the basal expression level of histidine decarboxylase mRNA of neurons located in the tuberomammillary nuclei of the posterior hypothalamus. The effects were both dose- and time-dependent. In conclusion, compensation of both static and dynamic deficits is subtended by long-term adaptive mechanisms that could be facilitated pharmacologically using betahistine dihydrochloride.

Glyceraldehyde-3-phosphate dehydrogenase is a GABAA receptor kinase linking glycolysis to neuronal inhibition.

Protein phosphorylation is crucial for regulating synaptic transmission. We describe a novel mechanism for the phosphorylation of the GABA(A) receptor, which mediates fast inhibition in the brain. A protein copurified and coimmunoprecipitated with the phosphorylated receptor alpha1 subunit; this receptor-associated protein was identified by purification and microsequencing as the key glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Molecular constructs demonstrated that GAPDH directly phosphorylates the long intracellular loop of GABA(A) receptor alpha1 subunit at identified serine and threonine residues. GAPDH and the alpha1 subunit were found to be colocalized at the neuronal plasma membrane. In keeping with the GAPDH/GABA(A) receptor molecular association, glycolytic ATP produced locally at plasma membranes was consumed for this alpha1 subunit phosphorylation, possibly within a single macrocomplex. The membrane-attached GAPDH is thus a dual-purpose enzyme, a glycolytic dehydrogenase, and a receptor-associated kinase. In acutely dissociated cortical neurons, the rundown of the GABA(A) responses was essentially attributable to a Mg(2+)-dependent phosphatase activity, which was sensitive to vanadate but insensitive to okadaic acid or fluoride. Rundown was significantly reduced by the addition of GAPDH or its reduced cofactor NADH and nearly abolished by the addition of its substrate glyceraldehyde-3-phosphate (G3P). The prevention of rundown by G3P was abolished by iodoacetamide, an inhibitor of the dehydrogenase activity of GAPDH, indicating that the GABA(A) responses are maintained by a glycolysis-dependent phosphorylation. Our results provide a molecular mechanism for the direct involvement of glycolysis in neurotransmission.

Betahistine dihydrochloride interaction with the histaminergic system in the cat: neurochemical and molecular mechanisms.

Drugs interfering with the histaminergic system facilitate behavioral recovery after vestibular lesion, likely by increasing histamine turnover and release. The effects of betahistine (structural analogue of histamine) on the histaminergic system were tested by quantifying messenger RNA for histidine decarboxylase (enzyme synthesizing histamine) by in situ hybridization and binding to histamine H(3) receptors (mediating, namely, histamine autoinhibition) using a histamine H(3) receptor agonist ([(3)H]N-alpha-methylhistamine) and radioautography methods. Experiments were done in brain sections of control cats (N=6) and cats treated with betahistine for 1 (N=6) or 3 (N=6) weeks. Betahistine treatment induced symmetrical changes with up-regulation of histidine decarboxylase mRNA in the tuberomammillary nucleus and reduction of [(3)H]N-alpha-methylhistamine labeling in both the tuberomammillary nucleus, the vestibular nuclei complex and nuclei of the inferior olive. These findings suggest that betahistine upregulates histamine turnover and release, very likely by blocking presynaptic histamine H(3) receptors, and induces histamine H(3) receptor downregulation. This action on the histaminergic system could explain the effectiveness of betahistine in the treatment of vertigo and vestibular disease.

Immunohistological localization of the myelinating cell-specific receptor LP(A1).

LP(A1) (also termed Edg-2 or VZG-1) is a G-protein-coupled receptor for lysophosphatidic acid and its gene transcripts have been found selectively expressed by mature myelin-producing cells. We have raised in rabbit a polyclonal antibody against a sequence unique to LP(A1) and common to rat, mouse, and human orthologues. In Western blots, LP(A1) immunoreactivity appeared as 44-53 kDa bands in extracts from recombinant RH7777 cells expressing LP(A1), mouse purified oligodendrocytes, or human white matter, but not from wild-type RH7777 cells or purified astrocytes. In glial cultures, LP(A1) immunoreactivity was restricted to oligodendrocytes, appeared at cell membrane and processes, colocalized with myelin basic protein, and appeared before myelin/oligodendrocyte glycoprotein. In slices of rat and human brains, LP(A1) immunoreactivity was found in myelinated tracts, as well as in oligodendrocyte somata and their myelinating fibers. Immunoreactivities of LP(A1) and myelin basic protein colocalized in the brain, but oligodendrocyte soma showed stronger signals for LP(A1) than myelinated fibers, whereas the reverse was true for myelin basic protein. These results strengthen the view that LP(A1) is involved in myelin formation or maintenance.