The NKCC1 antagonist bumetanide mitigates interneuronopathy associated with ethanol exposure in utero
- Geisel School of Medicine at Dartmouth, United States
- University of Minnesota Twin Cities, United States
Figures
Figure 1 with 2 supplements
Ethanol induces a depolarizing shift in GABA reversal potential in embryonic MGE-derived GABAergic cortical interneurons that is normalized by the NKCC1 inhibitor bumetanide.
(a) Hoffman modulated contrast image of acute telencephalic slice of E14.5 mouse brain showing patch clamp electrode (green outline) holding a tdTomato fluorescent Nkx2.1+ MGE-derived GABAergic …
Figure 1—figure supplement 1
50 mM ethanol induces a depolarizing shift in GABA reversal potential in embryonic MGE-derived GABAergic cortical interneurons.
(a) I/V plot of peak GABA-induced current over holding potential defines EGABA as the x-intercept under control (aCSF; black) and ethanol exposure (EtOH; red) conditions. Dotted blue line denotes …
Figure 1—figure supplement 2
I/V plot of peak GABA-induced current over holding potential in bumetanide pre-treated cells.
I/V plot of peak GABA induced current over holding potential defines EGABA as the x-intercept under bumetanide control (Bumet; black) and 6.5 mM ethanol exposure (EtOH; red) conditions. Dotted blue …
Figure 2 with 2 supplements
Bumetanide attenuates ethanol-induced potentiation of depolarizing GABA responses in embryonic MGE-derived GABAergic cortical interneurons.
(a) Peak current amplitude recorded from Nkx2.1+ MGE-derived GABAergic interneurons in slices of E14.5 mouse telencephalon in response to focal application of GABA (50 µM) under control (aCSF; …
Figure 2—figure supplement 1
50 mM ethanol induces a potentiation of depolarizing GABA responses in embryonic MGE-derived GABAergic cortical interneurons.
Peak current amplitude recorded from Nkx2.1+ MGE-derived GABAergic interneurons in slices of E14.5 mouse telencephalon in response to focal application of GABA (50 µM) under control (aCSF; black) …
Figure 2—figure supplement 2
Bumetanide treatment does not change the GABA response amplitude.
The mean GABA response amplitude in control (aCSF) and bumetanide pre-treated (Bumet) cells. Numbers above x-axis denote number of litters and number of cells recorded in (). Unpaired t-test.
Figure 3
Co-treatment with bumetanide prevents ethanol’s enhancement of tangential migration in vitro.
(a) Fluorescent images of organotypic E14.5 Nkx2.1-Cre/Ai14 mouse telencephalic brain slices treated with vehicle, ethanol (50 mM) + vehicle (EtOH + Vehicle), ethanol + bumetanide (EtOH+Bumet, 20 …
Figure 4
Maternal bumetanide treatment prevents ethanol-induced escalation of tangential migration in vivo.
Nkx2.1+ cells were identified by tdTomato fluorescence and quantified in the dorso-medial telencephalon of E16.5 embryonic brain. (a) Fluorescent images counterstained with DAPI following control, …
Figure 5
Treatment of binge ethanol-exposed dams with bumetanide prevents the interneuronopathy associated with ethanol exposure in utero.
(a) Histological sections of young adult mouse prefrontal cortex processed for parvalbumin immunofluorescence, counterstained with DAPI, and binned by functional region and layer using DAPI …
Figure 6
Maternal bumetanide treatment prevents the deficits in behavioral flexibility seen with ethanol exposure in utero.
(a) Mean number of errors committed in the modified Barnes maze by young adult mice born to control, ethanol consuming (EtOH) and ethanol consuming plus bumetanide treated (EtOH+Bumet) dams across …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Genetic reagent (M. musculus) | Nkx2.1-Cre | The Jackson Laboratory | JAX: 00861 MGI: J:131144 | PMID: 17990269 |
| Genetic reagent (M. musculus) | Ai14 | The Jackson Laboratory | JAX: 007914 MGI: J:155793 | PMID: 20023653 |
| Antibody | Anti-parvalbumin (mouse polyclonal) | Millipore Sigma | Cat#: MAB 1572 | 1:1000 |
| Chemical compound, drug | GABA | Millipore Sigma | Cat#: A2129 | |
| Chemical compound, drug | Bumetanide | Millipore Sigma | Cat#: B-3023 | |
| Software, algorithm | ImageJ | NIH | https://imagej.nih.gov/ij/ | |
| Software, algorithm | Adobe Photoshop | Adobe Systems, San Jose, CA | http://www.adobe.com/products/photoshop.html | |
| Software, algorithm | Graphpad Prism | Graphpad Software, Inc, La Jolla, CA | http://www.graphpad.com/ | |
| Software, algorithm | G*Power 3.1 | Heinrich Heine University Düsseldorf | RRID: SCR_013726 |
Additional files
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Transparent reporting form
- https://doi.org/10.7554/eLife.48648.012
