Exploration of beta-arrestin isoform signaling pathways in delta opioid receptor agonist-induced convulsions.

The δ-opioid receptor (δOR) has been considered as a therapeutic target in multiple neurological and neuropsychiatric disorders particularly as δOR agonists are deemed safer alternatives relative to the more abuse-liable µ-opioid receptor drugs. Clinical development of δOR agonists, however, has been challenging in part due to the seizure-inducing effects of certain δOR agonists. Especially agonists that resemble the δOR-selective agonist SNC80 have well-established convulsive activity. Close inspection suggests that many of those seizurogenic δOR agonists efficaciously recruit ß-arrestin, yet surprisingly, SNC80 displays enhanced seizure activity in ß-arrestin 1 knockout mice. This finding led us to hypothesize that perhaps ß-arrestin 1 is protective against, whereas ß-arrestin 2 is detrimental for δOR-agonist-induced seizures. To investigate our hypothesis, we characterized three different δOR agonists (SNC80, ADL5859, ARM390) in cellular assays and in vivo in wild-type and ß-arrestin 1 and ß-arrestin 2 knockout mice for seizure activity. We also investigated downstream kinases associated with ß-arrestin-dependent signal transduction. We discovered that δOR agonist-induced seizure activity strongly and positively correlates with ß-arrestin 2 efficacy for the agonist, but that indirect inhibition of ERK activation using the MEK inhibitor SL327 did not inhibit seizure potency and duration. Inhibition of the PI3K/AKT/mTOR signaling with honokiol but not PQR530, attenuated SNC80 seizure duration in ß-arrestin 1 knockout, but honokiol did not reduce SNC80-induced seizures in wild-type mice. Ultimately, our results indicate that ß-arrestin 2 is correlated with δOR agonist-induced seizure intensity, but that global ß-arrestin 1 knockout mice are a poor model system to investigate their mechanism of action.

Generation and basic characterization of a gene-trap knockout mouse model of Scn2a with a substantial reduction of voltage-gated sodium channel Nav 1.2 expression.

Large-scale genetic studies revealed SCN2A as one of the most frequently mutated genes in patients with neurodevelopmental disorders. SCN2A encodes for the voltage-gated sodium channel isoform 1.2 (Nav 1.2) expressed in the neurons of the central nervous system. Homozygous knockout (null) of Scn2a in mice is perinatal lethal, whereas heterozygous knockout of Scn2a (Scn2a+/- ) results in mild behavior abnormalities. The Nav 1.2 expression level in Scn2a+/- mice is reported to be around 50-60% of the wild-type (WT) level, which indicates that a close to 50% reduction of Nav 1.2 expression may not be sufficient to lead to major behavioral phenotypes in mice. To overcome this barrier, we characterized a novel mouse model of severe Scn2a deficiency using a targeted gene-trap knockout (gtKO) strategy. This approach produces viable homozygous mice (Scn2agtKO/gtKO ) that can survive to adulthood, with about a quarter of Nav 1.2 expression compared to WT mice. Innate behaviors like nesting and mating were profoundly disrupted in Scn2agtKO/gtKO mice. Notably, Scn2agtKO/gtKO mice have a significantly decreased center duration compared to WT in the open field test, suggesting anxiety-like behaviors in a novel, open space. These mice also have decreased thermal and cold tolerance. Additionally, Scn2agtKO/gtKO mice have increased fix-pattern exploration in the novel object exploration test and a slight increase in grooming, indicating a detectable level of repetitive behaviors. They bury little to no marbles and have decreased interaction with novel objects. These Scn2a gene-trap knockout mice thus provide a unique model to study pathophysiology associated with severe Scn2a deficiency.