Effects of Gαi 2 and Gαz protein knockdown on alpha 2A -adrenergic and cannabinoid CB 1 receptor regulation of MEK-ERK and FADD pathways in mouse cerebral cortex.

Background: Alpha _2A -adrenergic (α _2A -AR) and cannabinoid CB ₁ (CB ₁ -R) receptors exert their functions modulating multiple signaling pathways, including MEK-ERK (extracellular signal-regulated kinases) and FADD (Fas-associated protein with death domain) cascades. These molecules are relevant in finding biased agonists with fewer side effects, but the mechanisms involving their modulations by α _2A -AR- and CB ₁ -R in vivo are unclear. This study investigated the roles of Gαi ₂ and Gαz proteins in mediating α _2A -AR- and CB ₁ -R-induced alterations of MEK-ERK and FADD phosphorylation (p-) in mouse brain cortex.
Methods: Gαi ₂ or Gαz protein knockdown was induced in mice with selective antisense oligodeoxinucleotides (ODNs; 3 nmol/day, 5 days) prior to UK-14,304 (UK or brimonidine; 1 mg/kg) or WIN55212-2 (WIN; 8 mg/kg) acute treatments. Inactivated (p-T ²⁸⁶ ) MEK1, activated (p-S ^217/221 ) MEK1/2, activated (p-T ²⁰² /Y ²⁰⁴ ) ERK1/2, p-S ¹⁹¹ FADD, and the corresponding total forms of these proteins were quantified by immunoblotting.
Results: Increased (+ 88%) p-T ²⁸⁶ MEK1 cortical density, with a concomitant reduction (-43%) of activated ERK was observed in UK-treated mice. Both effects were attenuated by Gαi ₂ or Gαz antisense ODNs. Contrastingly, WIN induced Gαi ₂ - and Gαz-independent upregulations of p-T ²⁸⁶ MEK1 (+ 63%), p-S ^217/221 MEK1/2 (+ 86%), and activated ERK (+ 111%) in brain. Pro-apoptotic FADD was downregulated (- 34 to 39%) following UK and WIN administration, whereas the neuroprotective p-S ¹⁹¹ FADD was increased (+ 74%) in WIN-treated mice only. None of these latter effects required from Gαi ₂ or Gαz protein integrity.
Conclusion: The results indicate that α _2A -AR (UK), but not CB ₁ -R (WIN), agonists use Gαi ₂ and Gαz proteins to modulate MEK-ERK, but not FADD, pathway in mouse brain cortex.
(© 2021. Maj Institute of Pharmacology Polish Academy of Sciences.)