The NKCC1 Antagonist Bumetanide, Administered After Prenatal Alcohol Exposure, Mitigates Ethanol-Induced Aberrance in Tangential Migration of GABAergic Interneurons in Mouse Medial Prefrontal Cortex.

BACKGROUND: Prenatal ethanol (alcohol) exposure (PreEE) results in abnormal tangential migration of medial ganglionic eminence (MGE)-derived GABAergic interneurons (GINs) in the embryonic mouse medial prefrontal cortex (mPFC); later in life, this is associated with impaired behavioral flexibility commonly seen in fetal alcohol spectrum disorders (FASD). The PreEE-induced abnormal tangential migration involves, in part, ethanol potentiating GABAA receptor-mediated depolarization, the latter due to elevated intracellular chloride sustained primarily by the Na-K-Cl cotransporter NKCC1 in embryonic GINs. We previously reported that coincident administration of the NKCC1 antagonist bumetanide with PreEE prevents the PreEE-induced aberrant migration of GINs in the mPFC. However, such simultaneous dispensing rarely occurs in real life. In this light, the present study assessed whether bumetanide administered after PreEE mitigates the PreEE-induced aberrant tangential migration.

METHODS: NKCC1-like immunofluorescence was assessed following immunohistochemical processing of 30-μm cryosections derived from embryonic day (E)14.5 wild-type mouse brains. Nkx2.1Cre-Ai14 mice were used to genetically label MGE-derived GINs. Pregnant dams were fed 5% ethanol (w/w) from E13.5 to E16.5, followed by bumetanide treatment (320 mg/kg liquid diet) from E17.5 until postnatal day (P) 0 (birth). Forty-micron cryosections from brains of progeny were collected at P0, and the density as well as spatial distribution of tdTomato-fluorescent (Nkx2.1+) GINs in the mPFC quantified.

RESULTS: Within the timeframe of PreEE, the NKCC1 protein is expressed ubiquitously in the embryonic mouse brain, including the dorsomedial telencephalon, the precursor to mPFC. PreEE increased the overall density of Nkx2.1+ GINs and accelerated their dorsal-to-ventral dispersion across the developing mPFC. Post-PreEE treatment with the NKCC1 antagonist bumetanide diminished these effects, reducing dorsal accumulation while preserving the baseline population of GINs.

CONCLUSIONS: These results indicate that blocking the NKCC1 chloride importer bumetanide can mitigate PreEE-induced interneuronopathy when administered after PreEE, implicating NKCC1 as a potential pharmacological target for post-PreEE therapeutic intervention of FASD.