Suppressed kindling epileptogenesis in mice with ectopic overexpression of galanin

Abstract

The neuropeptide galanin has been shown to suppress epileptic seizures. In cortical and hippocampal areas, galanin is normally mainly expressed in noradrenergic afferents. We have generated a mouse overexpressing galanin in neurons under the plateletderived growth factor B promoter. RIA and HPLC analysis revealed up to 8-fold higher levels of galanin in transgenic as compared with wild-type mice. Ectopic galanin overexpression was detected especially in dentate granule cells and hippocampal and cortical pyramidal neurons. Galanin-overexpressing mice showed retardation of seizure generalization during hippocampal kindling, a model for human complex partial epilepsy. The high levels of galanin in mossy fibers found in the transgenic mice were further increased after seizures. Frequency facilitation of field excitatory postsynaptic potentials, a form of short-term synaptic plasticity assessed in hippocampal slices, was reduced in mossy fiber-CA3 cell synapses of galanin-overexpressing mice, indicating suppressed glutamate release. This effect was reversed by application of the putative galanin receptor antagonist M35. These data provide evidence that ectopically overexpressed galanin can be released and dampen the development of epilepsy by means of receptor-mediated action, at least partly by reducing glutamate release from mossy fibers.G alanin, a 29-aa neuropeptide (1), is widely distributed throughout the rat brain (2, 3) and coexists with classic transmitters in several systems (4), with particularly high levels in noradrenergic locus coeruleus neurons (5, 6). Galanin-binding sites have been mapped (7,8), and three receptor subtypes, GalR1 (9-11), 49), and GalR3 (15), have been cloned. There is evidence for involvement of galanin in multiple neuronal functions (16,17), as also revealed in recent studies on transgenic mice (18)(19)(20).Many studies support an important role of neuropeptides in seizure activity (21)(22)(23). This role is true also for galanin. Thus, intrahippocampal injection of galanin during status epilepticus (SE) in rats has powerful anticonvulsant effects (24). In accordance with these data, mice overexpressing the galanin gene under the dopamine-␤-hydroxylase promoter, presumably in noradrenergic fibers, show increased resistance to SE and kainate-and pentylenetetrazol-evoked seizures (25). Conversely, administration of galanin antagonists, such as M35 (26), and targeted disruption of the galanin gene increase seizure susceptibility in these models (24,25). Taken together, available data indicate that galanin released from noradrenergic fibers (3, 5, 27) can suppress epileptic seizures. Although the underlying mechanisms are not known, it has been shown that galanin can inhibit glutamate release by means of activation of presynaptic ATP-dependent K ϩ channels (28).We speculated that galanin release from neurons not normally containing this neuropeptide, e.g., in the epileptic focus, might be necessary to optimize the seizure-suppressant action of galanin. To explore the functional conseque...

Galanin expressed in the excitatory fibers attenuates synaptic strength and generalized seizures in the piriform cortex of mice

“…Further incubation with Streptavidine-HRP, biotinyl tyramide reagent and Alexa-488 fluorophore finalized the reaction. The specificity of antibody binding was confirmed by omission 11 of the primary antibody on a subset of sections (Kokaia, M., Holmberg, K., et al 2001;Kuteeva, E., Calza, L., et al 2004).…”

“…GalOE mice were generated as described earlier (Kokaia, M., Holmberg, K., et al 2001), and the transgenic status of animals was confirmed by PCR (Kokaia, M., Holmberg, K., et al 2001). All experiments were performed according to National Institutes of Health guidelines and approved by the local Ethical Committee.…”

“…In line with these observations, galanin gene knockout mice were shown to be more prone to develop SE seizures, while transgenic mice overexpressing the galanin gene in noradrenergic neurons under the dopamine β-hydroxylase promoter were more resistant to the induction of SE (Mazarati, A.M., Hohmann, J.G., et al 2000). Earlier, we have demonstrated that even ectopic galanin overexpression under the PDGF-B promotor suppressed hippocampal kindling 6 epileptogenesis in mice, and was associated with attenuated frequency facilitation in the mossy fiber -CA3 cell synapses (Kokaia, M., Holmberg, K., et al 2001). …”

“…Similarly to other neuropeptides such as NPY (Vezzani, A. and Sperk, G. 2004), these seizure-induced changes in galanin expression have been taken to suggest that galanin is involved in seizure regulation, presumably exerting a protective action. Indeed, accumulating evidence indicates that galanin can modulate seizure activity in the brain, as shown in different animal models of epilepsy (Kokaia, M., Holmberg, K., et al 2001;Mazarati, A.M. 2004;Mazarati, A.M., Hohmann, J.G., et al 2000;Mazarati, A.M., Liu, H., et al 1998;Mazarati, A.M., Lu, X., et al 2004b;Mazarati, A.M. and Wasterlain, C.G. 2002;McColl, C.D., Jacoby, A.S., et al 2006).…”

“…We particularly chose the synapses formed by OB mitral cell axons onto the pyramidal cells of the PC, because (1) galanin is ectopically expressed in the OB mitral cell bodies and axons of galanin overexpressing (GalOE) mice (Kokaia, M., Holmberg, K., et al 2001;Kuteeva, E., Calza, L., et al 2004), and (2) these synapses can play an important role in the spread of epileptic seizures from the OB.…”