Activation of protease-activated receptor 2 induces hyperexcitability in human sensory neurons

Editor’s note: The research described below comes from a recipient of a 2025 MSC Travel Grant supporting travel to the 67th Annual Scientific Meeting of the American Headache Society. These grants reimburse travel expenses for those who have had their abstract for a presentation or poster accepted at a meeting.

By Mandee Schaub, PhD student, University of Texas at Dallas, US.

What is the research gap that your study addresses?

Migraine is the 2nd leading cause of disability worldwide and currently available therapeutics are only 50% effective in 50% of patients. We are trying to validate a mechanistic contributor to migraine as a therapeutic target.

What is your research hypothesis?

We hypothesize that activation of protease-activated receptor 2 contributes to increased sensory neuron excitability in human sensory neurons.

What methodology did you use to address your research hypothesis?

We used whole-cell patch-clamp electrophysiology to assess functional properties of neurons cultured from dorsal root ganglion (hDRG) of human organ donors. We assessed hDRG cells for spontaneous activity, rheobase, and evoked action potential firing to both ramp and step stimuli. hDRGs were collected from organ donors, cultured, and patched on days 3-5 in vitro. We treated neurons with both selective PAR2 activator 2-AT and physiological PAR2 activators elastase and trypsin to assess PAR2 activation induced- hyperexcitability in hDRG neurons. We then used selective PAR2 antagonist C937 to block excitability produced by the agonists.

What are the main results of your study?

Selective PAR2 activation results in hyperexcitability of hDRG neurons. We saw an increase in spontaneous activity, decrease in rheobase (lowest stimulus needed to elicit an action potential) and increase in number of action potentials fired in response to a ramp stimulus. We then saw an increase in excitability produced by treatment with both elastase and trypsin, and these increases could be partially blocked by pre-treatment with a PAR2 antagonist, C937.

What conclusions did you reach based on your results?

PAR2 activation does cause hyperexcitability of human sensory neurons and that hyperexcitability can be blocked with PAR2 antagonism.

What are the limitations of your study?

There are many types of neurons in the DRG, so we don’t know what subtypes we are patching. There may be a larger effect in a different subtype of neurons. In addition, because we are culturing the neurons, there may be subtypes that are not included in our samples because they may not live through the culture process. The cells we are using are not from migraine-relevant tissue, but tissue relevant for body pain. In the future we hope to repeat these experiments in human trigeminal ganglion cells, which would be more directly relevant for migraine.

What is the relevance of your study to migraine?

This study investigates the direct actions of a known migraine target in a human cell type very similar to those that transmit migraine pain signals. Further, the physiological stimuli used, elastase and trypsin, are released from immune cells in response to inflammation or injury, and likely during a migraine attack. The trigeminal ganglion neurons (TG) that innervate the dura are closely associated with immune cells that release proteases that activate PAR2, like those we investigated in this study. This is preliminary work that will lead to further investigation in hTG cells, which will be the most migraine-relevant study.