Knockdown of Tlr3 in dorsal striatum reduces ethanol consumption and acute functional tolerance in male mice

• Published in: Brain, behavior, and immunity Vol. 118; pp. 437 - 448
• Main Authors: Dilly, Geoffrey A., Blednov, Yuri A., Warden, Anna S., Ezerskiy, Lubov, Fleischer, Caleb, Plotkin, Jesse D., Patil, Shruti, Osterndorff-Kahanek, Elizabeth A., Mayfield, Jody, Mayfield, R. Dayne, Homanics, Gregg E., Messing, Robert O.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier Inc 01.05.2024
• Subjects: Acute functional tolerance to ethanol; Alcohol Drinking - metabolism; Animals; Diazepam; Ethanol - pharmacology; Male; Mice; Mice, Inbred C57BL; Poly I-C - pharmacology; Poly(I:C); Rotarod ataxia; Signal Transduction; Tlr3 knockdown in dorsal striatum; Tlr3F/F mice; Toll-Like Receptor 3 - metabolism; Two-bottle choice every-other-day ethanol consumption; Rotarod ataxia; Acute functional tolerance to ethanol; Poly(I:C); Tlr3F/F mice; Two-bottle choice every-other-day ethanol consumption; Diazepam; Tlr3 knockdown in dorsal striatum; Tlr3(F/F) mice
• ISSN: 0889-1591; 1090-2139
• DOI: 10.1016/j.bbi.2024.03.021

•Tlr3 floxed (Tlr3F/F) mice were generated using CRISPR/Cas9 genome editing.•Cre recombination knocked down Tlr3 mainly in neurons.•Knockdown in dorsal striatum (DS) ↑ ataxia by ethanol and ↓ ethanol consumption.•Neuronal TLR3 signaling within the DS regulates ethanol drinking and intoxication. Systemic activation of toll-like receptor 3 (TLR3) signaling using poly(I:C), a TLR3 agonist, drives ethanol consumption in several rodent models, while global knockout of Tlr3 reduces drinking in C57BL/6J male mice. To determine if brain TLR3 pathways are involved in drinking behavior, we used CRISPR/Cas9 genome editing to generate a Tlr3 floxed (Tlr3F/F) mouse line. After sequence confirmation and functional validation of Tlr3 brain transcripts, we injected Tlr3F/F male mice with an adeno-associated virus expressing Cre recombinase (AAV5-CMV-Cre-GFP) to knockdown Tlr3 in the medial prefrontal cortex, nucleus accumbens, or dorsal striatum (DS). Only Tlr3 knockdown in the DS decreased two-bottle choice, every-other-day (2BC-EOD) ethanol consumption. DS-specific deletion of Tlr3 also increased intoxication and prevented acute functional tolerance to ethanol. In contrast, poly(I:C)-induced activation of TLR3 signaling decreased intoxication in male C57BL/6J mice, consistent with its ability to increase 2BC-EOD ethanol consumption in these mice. We also found that TLR3 was highly colocalized with DS neurons. AAV5-Cre transfection occurred predominantly in neurons, but there was minimal transfection in astrocytes and microglia. Collectively, our previous and current studies show that activating or inhibiting TLR3 signaling produces opposite effects on acute responses to ethanol and on ethanol consumption. While previous studies, however, used global knockout or systemic TLR3 activation (which alter peripheral and brain innate immune responses), the current results provide new evidence that brain TLR3 signaling regulates ethanol drinking. We propose that activation of TLR3 signaling in DS neurons increases ethanol consumption and that a striatal TLR3 pathway is a potential target to reduce excessive drinking.