Connexin-36 Knock-Out Mice have Increased Threshold for Kindled Seizures: Role of GABA Inhibition

Abstract

Blockade of gap junctions (GJs) has been shown to reduce seizures in different epilepsy models. Gap junctionmediated, electrically-coupled neuronal networks have been implicated in neuronal synchronization, which is the hallmark of seizure activity. To further understand the role of GJs in seizures, in particular hippocampal seizures, we evaluated electrophysiological responses in the dentate gyrus subfield of the hippocampus, including GABA-mediated recurrent inhibition, seizuregenic stimulation, and seizure activity in response to perforant path kindling in Cx36 knockout (KO) mice compared to wild-type (WT) controls. In anesthetized mice, Cx36 KO mice were characterized by enhanced GABA-mediated recurrent inhibition. Stimulation of the perforant path at 10 Hz for 10 sec (i.e., 100 pulses) markedly enhanced population spike (PS) amplitudes and reduced paired-pulse GABA-mediated inhibition during the stimulation, induced an after discharge for approximately 10 sec at 7-10 sec into the stimulation, and suppressed PS amplitudes postictally for 5-10 min in both KO and WT mice. Repeated epochs of 10 Hz for 10 sec stimulation of the perforant path at 20 min intervals resulted in progressive and persistent disinhibition of paired-pulse responses in WT, but not KO mice. In freely-behaving seizure studies, stimulation of the perforant path (10 Hz for 10 sec) once/day resulted in progressive Stage I-IV seizures in both WT and KO mice. Once Stage IV was achieved, another Stage IV seizure could be elicited each day with the same behavioral response (i.e., Stage V). The threshold for kindled seizures was significantly greater in KO mice compared to WT controls for most stages of seizures. The Cx36 antagonist mefloquine (MFQ) and the typical anticonvulsant pentobarbital reduced Stage IV seizures in both WT and KO mice. Knock-out mice were more sensitive to the anti-epileptic effects of the typical anti-convulsant pentobarbital (20 mg/ kg). Taken together, these findings support the emerging view that reduction in GJ-mediated GABA electrical coupling reduces seizures, perhaps through enhancement of GABAergic feedback inhibition, which results from the uncoupling of the GABA recurrent interneurons from the resistive load that is inherent in their electrical connectivity.

Mefloquine-induced conformational shift in Cx36 N-terminal helix leading to channel closure mediated by lipid bilayer

“…Among them, MFQ, an FDA-approved antimalarial compound, is used for the prevention or treatment of mild and moderate malaria 20 , with neuropsychiatric side effects 14,18,21 . Later, it was discovered that MFQ specifically blocks Cx36 GJC, contributing to these side effects, such as spreading depression, NMDAR-mediated neuronal death, tremor, motor deficits, convulsant, stage IV seizures, cell death upon oxygenglucose deprivation, and hypersensitivity to pain [22][23][24][25][26][27][28][29] . Additionally, it is noteworthy that MFQ exhibits 10-to 100-fold higher sensitivity to Cx36 (IC50 value of 310 nM) and Cx50 (IC50 value of 1.1 μM) compared to other connexins.…”

Effects of Constitutive and Acute Connexin 36 Deficiency on Brain-Wide Susceptibility to PTZ-Induced Neuronal Hyperactivity

“…Previous work has examined the roles of Cx36 in the pathogenesis of seizures, but there has been no consensus on whether Cx36 increases or decreases seizure susceptibility (Gajda et al, 2005 ; Jacobson et al, 2010 ; Voss et al, 2010 ; Shin, 2013 ). Jacobson et al ( 2010 ) found that Cx36 knockout mice exhibited an increase in seizure-like behaviors following the administration pentylenetetrazol (PTZ; a GABA(A)-receptor antagonist), indicating that normal expression of Cx36 may be protective against seizure-inducing conditions.…”

Effects of constitutive and acute Connexin 36 deficiency on brain-wide susceptibility to PTZ-induced neuronal hyperactivity

“…Previous work has examined the roles of Cx36 in the pathogenesis of seizures, but there has been no consensus on whether Cx36 increases or decreases seizure susceptibility (Gajda, Szupera, Blazsó, & Szente, 2005;Jacobson et al, 2010;Shin, 2013;Voss, Mutsaerts, & Sleigh, 2010a). Jacobson et al (2010) found that Cx36 knockout mice exhibited an increase in seizurelike behaviors following the administration pentylenetetrazol (PTZ; a GABA(A)-receptor antagonist), indicating that normal expression of Cx36 may be protective against seizure-inducing conditions.…”