Prosper N'Gouemo

Targeting BK (big potassium) channels in epilepsy

Introduction: Epilepsies are disorders of neuronal excitability characterized by spontaneous and recurrent seizures. Ion channels are critical for regulating neuronal excitability and, therefore, can contribute significantly to epilepsy pathophysiology. In particular, large conductance, Ca2+-activated K+ (BKCa) channels play an important role in seizure etiology. These channels are activated by both membrane depolarization and increased intracellular Ca2+. This unique coupling of Ca2+ signaling to membrane depolarization is important in controlling neuronal hyperexcitability, as outward K+ current through BKCa channels hyperpolarizes neurons. Areas covered: BKCa channel structure–function and the role of these channels in epilepsy pathophysiology. Expert opinion: Loss-of-function BKCa channel mutations contribute to neuronal hyperexcitability that can lead to temporal lobe epilepsy, tonic–clonic seizures and alcohol withdrawal seizures. Similarly, BKCa channel blockade can trigger seizures and status epilepticus. Paradoxically, some mutations in BKCa channel subunit can give rise to channel gain-of-function that leads to development of idiopathic epilepsy (primarily absence epilepsy). Seizures themselves also enhance BKCa channel currents associated with neuronal hyperexcitability, and blocking BKCa channels suppresses generalized tonic–clonic seizures. Thus, both loss-of-function and gain-of-function BKCa channels might serve as molecular targets for drugs to suppress certain seizure phenotypes including temporal lobe seizures and absence seizures, respectively.

Amiloride delays the onset of pilocarpine-induced seizures in rats

Recent evidence suggests that amiloride, a potent and nonselective blocker of acid-sensing ion channels, suppresses generalized seizures induced by maximal electroshock and pentylenetrazole. Here I further determined and quantified the effects of amiloride on the occurrence of limbic seizures and status epilepticus-induced by intraperitoneal administration of pilocarpine, a muscarinic acetylcholine receptor agonist. Pretreatment with various doses (5, 10, 30, 100, and 200 mg/kg) of amiloride significantly delayed the onset of the first episode of limbic seizures and the occurrence of status epilepticus following administration of pilocarpine (380 mg/kg). At the dose of 100 and 200 mg/kg, amiloride suppressed limbic seizures in 33% of pilocarpine-treated animals and significantly reduced the seizure severity score in 67% of the remaining animals. These findings suggest that amiloride may modulate seizure generation and propagation, probably via mechanisms involving acid-sensing ion channels in the pilocarpine model of temporal lobe epilepsy.

Seizure susceptibility is associated with altered protein expression of voltage-gated calcium channel subunits in inferior colliculus neurons of the genetically epilepsy-prone rat.

The inferior colliculus (IC) is the consensus site for seizure initiation in the genetically epilepsy-prone rat (GEPR). We have previously reported that the current density of high threshold voltage-activated (HVA) calcium (Ca(2+)) channels was markedly enhanced in IC neurons of the GEPR-3 (moderate seizure severity substrain of the GEPR). The present study examines whether subunit protein levels of HVA Ca(2+) channels are altered in IC neurons that exhibit enhanced Ca(2+) current density. Quantification shows that the levels of protein expression of the Ca(2+) channel pore-forming alpha1D (L-type) and alpha1E subunits (R-type) were significantly increased in IC neurons of seizure-naive GEPR-3s (SN-GEPR-3s) compared to control Sprague-Dawley (SD) rats. Significant increases and decreases in the levels of protein expression of Ca(2+) channel regulatory beta3 and alpha2delta subunits occurred in IC neurons of SN-GEPR-3s compared to control SD rats, respectively. No changes occurred in the protein expression of Ca(2+) channel pore-forming alpha1A (P/Q-type), alpha1B (N-type) and alpha1C (L-type) subunits in IC neurons of SN-GEPR-3s compared to control SD rats. A single seizure selectively enhanced protein expression of Ca(2+) channel alpha1A subunits in IC neurons of GEPR-3s. Thus, up-regulation of Ca(2+) channel alpha1D and alpha1E subunits may represent the molecular mechanisms for the enhanced current density of L- and R-type of HVA Ca(2+) channels in IC neurons of the GEPR, and may contribute to the genetic basis of their enhanced seizure susceptibility. The up-regulation of Ca(2+) channel alpha1A subunits induced by seizures may contribute to the increasing IC neuronal excitability that results from repetitive seizures in the GEPR.

Blockade of the sodium calcium exchanger exhibits anticonvulsant activity in a pilocarpine model of acute seizures in rats

Recent evidence suggests that the sodium calcium exchanger (NCX) may contribute to the etiology of pentylenetetrazol-induced seizures. Here we further investigated the role of NCX in the etiology of seizures by quantifying the effects of KB-R7943 and SN-6, potent inhibitors of the reverse mode of NCX subtypes 3 (NCX3) and 1 (NCX1), respectively, on the occurrence of acute seizures and status epilepticus induced by intraperitoneal administration of pilocarpine, a muscarinic acetylcholine receptor agonist. Pretreatment with KB-R7943 significantly reduced the incidence of pilocarpine-induced seizures and status epilepticus in 22-56% of treated animals. In the remaining animals that exhibited seizures, KB-R7943 pretreatment delayed the onset of seizures and status epilepticus, and reduced seizure severity. Delayed onset of seizures and reduced seizure severity also were seen following pretreatment with SN-6. These findings suggest that altered NCX activity may contribute to the pathophysiology of pilocarpine-induced seizures and status epilepticus.

Calcium channel dysfunction in inferior colliculus neurons of the genetically epilepsy-prone rat.

Voltage-gated calcium (Ca(2+)) channels are thought to play an important role in epileptogenesis and seizure generation. Here, using the whole cell configuration of patch-clamp techniques, we report on the modifications of biophysical and pharmacological properties of high threshold voltage-activated Ca(2+) channel currents in inferior colliculus (IC) neurons of the genetically epilepsy-prone rats (GEPR-3s). Ca(2+) channel currents were measured by depolarizing pulses from a holding potential of - 80 mV using barium (Ba(2+)) as the charge carrier. We found that the current density of high threshold voltage-activated Ca(2+) channels was significantly larger in IC neurons of seizure-naive GEPR-3s compared to control Sprague-Dawley rats, and that seizure episodes further enhanced the current density in the GEPR-3s. The increased current density was reflected by both a - 20 mV shifts in channel activation and a 25% increase in the non-inactivating fraction of channels in seizure-naive GEPR-3s. Such changes were reduced by seizure episodes in the GEPR-3s. Pharmacological analysis of the current density suggests that upregulation of L-, N- and R-type of Ca(2+) channels may contribute to IC neuronal hyperexcitability that leads to seizure susceptibility in the GEPR-3s.

Protein expression of small conductance calcium-activated potassium channels is altered in inferior colliculus neurons of the genetically epilepsy-prone rat

The genetically epilepsy-prone rat (GEPR) exhibits inherited predisposition to sound stimuli-induced generalized tonic-clonic seizures (audiogenic reflex seizures) and is a valid model to study the physiopathology of epilepsy. In this model, the inferior colliculus (IC) exhibits enhanced neuronal firing that is critical in the initiation of reflex audiogenic seizures. The mechanisms underlying IC neuronal hyperexcitability that leads to seizure susceptibility are not as yet fully understood. The present report shows that the levels of protein expression of SK1 and SK3 subtypes of the small conductance Ca2+-activated K+ channels were significantly decreased, while SK2 channel proteins were increased in IC neurons of seizure-naive GEPR-3s (SN-GEPR-3), as compared to control Sprague-Dawley rats. No significant change was found in the expression of BK channel proteins in IC neurons of SN-GEPR-3s. Single episode of reflex audiogenic seizures in the GEPR-3s did not significantly alter the protein expression of SK1-3 and BK channels in IC neurons compared to SN-GEPR-3s. Thus, downregulation of SK1 and SK3 channels and upregulation of SK2 channels provide direct evidence that these Ca2+-activated K+ channels play important roles in IC neuronal hyperexcitability that leads to inherited seizure susceptibility in the GEPR.

Ethanol withdrawal is accompanied by downregulation of calcium channel alpha 1B subunit in rat inferior colliculus neurons

Ethanol withdrawal enhances the current density of calcium (Ca(2+)) channels in inferior colliculus (IC) neurons. The present report shows that ethanol withdrawal markedly enhanced the susceptibility to seizures as it decreased significantly the protein levels of alpha(1B) subunit associated with N-type Ca(2+) channel in IC neurons of animals not tested for seizures. Thus, remodeling of N-type Ca(2+) channels may play an important role in neuronal hyperexcitability that leads to ethanol withdrawal seizures.

Neuronal Networks in Epilepsy: Comparative Audiogenic Seizure Networks

Abstract Neuronal networks involved in the epilepsies have been widely studied. The epilepsies are central nervous system disorders characterized by abnormal firing patterns of neurons within neuronal networks that are specific to the type of epilepsy. Critical brain sites or “hubs” within these networks and a variety of network control mechanisms have been identified in the various experimental forms of epilepsy. These epilepsy networks include models of developmental epilepsy, absence epilepsy, kindling epilepsy, status epilepticus, and generalized human epilepsy. One of the problems with much of the network-related information in these epilepsy models and human data is that the onset of the seizure is unpredictable, so the actual network nucleus (hub) for seizure initiation and the propagation pathway of the seizure through the network nuclei are uncertain. However, there are several animal models and certain forms of human epilepsy that are triggered by external (sensory) stimuli, which allows seizure onset to be under experimental control (“reflex” epilepsy). In most models of reflex epilepsy, the seizures are audiogenic seizures (AGS) that are induced by acoustic stimuli. Genetic AGS models have been studied in great detail, which allow comparisons of network hubs and mechanisms. These models include genetically epilepsy-prone rats (GEPR-9s and GEPR-3s) derived from the Sprague–Dawley strain and the Wistar-derived Strasbourg and Brazilian audiogenic rats and Russian (Krushinski–Molodkina) rats. Susceptibility to AGS can also be induced, and these models include thyroid deficiency–induced, ischemia-induced, and ethanol withdrawal–induced AGS. The induced models exhibit similar behavioral patterns of AGS, and these induced models have shed further light on the common hubs and mechanisms of AGS. AGS network elements involve the lower brainstem auditory network up through the critical hub in the inferior colliculus (IC), which projects onto other brainstem sites, including the deep layers of the superior colliculus, the pontine reticular formation, the periaqueductal gray, and the substantia nigra reticulata. Projections from these latter sites to the spinal cord generate the convulsive behaviors via overactivation of normal midbrain locomotor networks. Each of the sites plays a sequential dominant role during the behavior changes that occur during the convulsion. Glutamatergic and GABAergic mechanisms in certain of these network hubs, particularly in the IC, have been shown to be critically involved in initiating network activation. In addition, certain sites in the forebrain, including the amygdala, are differentially implicated in the different genetic AGS models. These different forms of AGS exhibit similar, but not identical, patterns of convulsive behavior, which are mediated by analogous but not identical network sites. The similarities and differences in network sites and mechanisms discovered with this comparative approach may provide a template for improved understanding of the “variations on a theme” seen in many human brain disorders that exhibit overlapping spectra of symptoms but do not display identical behavioral patterns.

Physiological and Pathophysiological Expansion of Neuronal Networks

Abstract Neuronal networks undergo experience-based changes, resulting in long-lasting expansion of the network involving both functional and structural alterations. Network expansion commonly involves response changes in conditional multireceptive neurons and is seen in vivo. The mechanisms for these long-lasting changes include long-term potentiation, which mediates certain neuroplastic changes involved in network expansion. A prominent example of network expansion is seen in the naturally occurring forms of audiogenic seizures (AGS), which are generalized convulsive seizures in rodents that are induced by loud sounds. Periodic repetition of AGS (AGS kindling) results in an increased duration of seizures and emergent alterations of convulsive behaviors in five different forms of AGS in rats, including the genetically epilepsy-prone rats (GEPR-9s and GEPR-3s), the Strasbourg Wistar AGS-susceptible rats, and the Wistar audiogenic rat strain from Brazil. Network expansion sites and mechanisms have been explored using many techniques, including brain imaging, focal microinjection, chemical stimulation, and electrophysiological techniques (electroencephalography and action potentials) in awake, behaving rats. Specific structures in the brainstem are the only ones required to produce AGS prior to AGS kindling. After AGS kindling, the network expands to include thalamic, limbic, and cortical structures; this differs in its specific details among the different AGS models and results in major differences between GEPR-9s and the other models for reasons that require further research. Neuroplastic mechanisms for network expansion in AGS kindling include N-methyl- d -aspartate receptor-mediated molecular events, and changes in cyclic adenosine monophosphate. AGS kindling is an analog of repetitive seizures that occur in many epileptic patients, and these contribute to mood disorders and memory and cognitive defects, and the network expansion to limbic system structures may explain these comorbidities in patients. Identification of specific sites of change in these expanded networks may provide therapeutic targets for drugs to prevent network expansion. The network expansion mechanisms observed in AGS kindling may shed light on potential general mechanisms of network expansion, including those seen in chronic pain and affective and anxiety disorders.

Amiloride and SN‐6 Suppress Audiogenic Seizure Susceptibility in Genetically Epilepsy‐Prone Rats

We have recently reported that amiloride, a potent and nonselective blocker of acid‐sensing ion channels, prevents the development of pilocarpine‐induced seizures and status epilepticus. Amiloride is also known to suppress the activity of Na+/Ca2+ and Na+/H+ exchangers that have been implicated in the pathophysiology of seizures. Here, we evaluated the effects of amiloride, SN‐6 (a potent blocker of Na+/Ca2+ exchangers) and zoniporide (a potent blocker of Na+/H+ exchangers) on acoustically evoked seizures (audiogenic seizures, AGS) in genetically epilepsy‐prone rats (GEPR‐3s), a model of inherited generalized epilepsy.