Cdk5 modulates orofacial pain through P2X2/3 purinergic receptor-mediated signaling in trigeminal neurons

Cyclin-dependent kinase 5 (Cdk5) plays a critical role in pain transmission by phosphorylating key nociceptive proteins. One of its recently identified substrates, the P2X2 subunit, is part of the P2X2/3 purinergic receptor (P2X2/3R), which is implicated in nociceptive signaling. However, the specific contribution of Cdk5 to purinergic receptor-mediated pain responses remains unclear. This study aimed to investigate the role of Cdk5 in pain responses elicited by α,β-methylene ATP (α,β-meATP), an agonist of P2X2/3R and P2X3R.

Both pharmacological inhibition and genetic loss-of-function approaches were used to assess the role of Cdk5. Calcium (Ca²⁺) imaging was performed in cultured murine trigeminal ganglion (TG) neurons to characterize α,β-meATP-evoked responses. Surface biotinylation assays in HEK293 cells were used to assess P2X2 receptor membrane expression. In vivo, facial pain was induced by injecting α,β-meATP into the whisker pad of mice. Behavioral responses were evaluated in conditional Cdk5 knockout mice targeting sensory neurons and in control littermates.

In TG neuron cultures, α,β-meATP evoked both fast and slow Ca²⁺ transients, which were sensitive to P2X3 or P2X2/3 channel blockade. Inhibition or genetic loss-of-function of Cdk5 predominantly accelerated the decay of slow responses, with minimal effects on fast responses. In HEK293 cells, Cdk5 activation did not alter the surface expression of P2X2 receptors. In vivo, α,β-meATP induced facial pain-like behaviors, including grooming and head flinching, which were significantly reduced in conditional Cdk5 knockout mice compared to controls.

These findings indicate that Cdk5 modulates orofacial pain by regulating purinergic receptor-mediated signaling in trigeminal neurons. Specifically, Cdk5 appears to influence the functional properties of P2X2-containing channels, most likely P2X2/3 heteromeric receptors, without affecting their membrane expression. This modulation of receptor kinetics may underlie the observed changes in nociceptive behavior, highlighting Cdk5 and P2X receptors as a potential target for pain modulation.