Valorization of 2-aminoethoxydiphenyl borate as a neurotherapeutic agent: modulation of calcineurin, TRPC1 and MARK2/GSK-3β signaling in scopolamine-triggered tauopathy.

In Alzheimer's disease (AD), amyloid beta (Aβ) plaques and hyperphosphorylated tau tangles drive neurodegeneration and cognitive decline. Disrupted calcium homeostasis-particularly via transient receptor potential canonical-1 (TRPC1) channels-contributes to tauopathy and disease progression. This study investigated the therapeutic potential of 2-aminoethoxydiphenyl borate (2-APB), a modulator of TRP channels, in a scopolamine-induced rat model of AD. Adult Wistar rats were assigned to six groups: normal control, disease control (scopolamine 2.5 mg/kg), three 2-APB treatment groups (2.5, 5.0 and 10.0 mg/kg) and a donepezil group (5.0 mg/kg), each with six animals. Treatments lasted three weeks. Cognitive performance was assessed using the Morris water maze, memory consolidation and open field tests. Biochemical assays measured acetylcholinesterase (AChE), calcineurin, oxidative stress markers (GSH, MDA) and inflammatory cytokines (TNF-α, IL-6, IL-1β). Protein and gene expression analyses (Western blot, qRT-PCR) evaluated tau, GSK-3β, TRPC1, MARK2, and calcineurin A/PPP3CA, alongside histopathological and immunohistochemical studies. 2-APB treatment significantly reduced pro-inflammatory cytokines, alleviated neuroinflammation, and decreased oxidative stress. Cognitive function improved, correlating with normalized AChE activity and preserved neuronal structure. At the molecular level, 2-APB reduced tau hyperphosphorylation at Ser396 and Thr231, likely through suppression of TRPC1-associated calcium entry and consequent modulation of calcium-dependent kinases (MARK2) and phosphatases (calcineurin), as well as the GSK-3β pathway. However, given that 2-APB is a broad-spectrum calcium signaling modulator - known to act on IP₃ receptors, SOCE/ORAI pathways, and multiple TRPC subtypes in addition to TRPC1-the present findings provide supportive evidence linking TRPC1 modulation to the observed neuroprotective effects. Although, the involvement of additional interconnected signaling mechanisms cannot be excluded and these findings should be interpreted within the context of 2-APB's pleiotropic pharmacological profile. Downregulation of TRPC1 and calcineurin A/PPP3CA gene expression supported 2-APB's role in restoring calcium balance and mitigating tau pathology. These results highlight 2-APB's potential in addressing key AD features-tauopathy, oxidative stress, neuroinflammation and cognitive impairment-warranting further preclinical and clinical research as a potential therapeutic for AD and related tauopathies.