Abstract
Modeling inflammatory injury via high-intensity stimulation of peripheral afferents at low frequencies (LFS) potentiates excitatory input onto spinal neurons projecting (PNs) to the midbrain periaqueductal gray (PAG), a region critical to pain processing. Therefore, we sought to identify the underlying afferent population driving this potentiation and how this enhanced synaptic drive might affect spino-PAG PN output to supraspinal pain circuits. We used 3–6-week-old male and female mice expressing Cre recombinase driven by expression of the TRPV1 gene, which is expressed in the lineage of virtually all C fibers, and Cre-dependent channelrhodopsin to stimulate C fibers. Spinal PNs were retrogradely labeled from the PAG using diI. We performed whole-cell patch-clamp recordings in spino-PAG PNs using a semi-intact spinal cord preparation. Stimulation of TRPV1-expressing (TRPV1+) peripheral afferents induced burst firing in virtually all spino-PAG PNs sampled. Inflammation increases the firing rates of C fibers to 1-2 Hz; therefore, we used LFS (1 ms, 470 nm LED pulses at 2 Hz for 2 mins) to model inflammatory injury. LFS induced a persistent (≥20 min) increase in the number of action potentials (APs) within a C fiber-induced burst (n=10, p=0.029), rise in the frequency of synaptically evoked after discharge (n=10, p=0.0004), and decrease in intraburst afterhyperpolarization (n=10, p=0.037). Furthermore, LFS induced a transient membrane depolarization (Mean=6.796 mV, n=12, p<0.0001). Additional experiments suggested the enhanced synaptic excitability relied on postsynaptic G protein-coupled signaling, NMDA receptors, and TRPV1+ afferent input. Finally, LFS may increase the intrinsic excitability of spino-PAG PNs—by decreasing AP threshold (n=14, p=0.015) and increasing membrane resistance (n=14, p=0.0003). LFS had no effect on the membrane potential nor the firing frequency of spino-PAG PNs. In summary, this work suggests that LFS of TRPV1+ fibers persistently enhances spino-PAG projection neuron output, likely leading to a lasting increase in the activation of PAG neural circuits. Grant support from F32NS123008 NINDS; Brewer, C. B. T32DA035165 NIDA; Mackey, S. C. R01DA011289 NIDA; Kauer, J. A.
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