Larry L. Bellinger

Purpose: It has become more apparent that studies focusing on the effect of sex steroids on orofacial pain in humans and animals require careful attention to the methods of hormone replacement. As a result, the purpose of this review is to highlight differences in estrogen concentration and its effects on temporomandibular joint (TMJ) pain and behavior.Methods: This literature review searched keywords TMJ, orofacial, pain, nociception sex steroids, estrous cycle, menstrual cycle estradiol, and progesterone.Results: In the case of estrogen, the change in concentration was a consideration, as a rise in estrogen can lead to protective effects and a decrease in estrogen appears to exacerbate the pain response. The dosage and timing was important when administering sex steroids as many orofacial pain studies conducted on animals have used large pharmacologic dosages and/or have given the hormone in a nonphysiologic way that does not mimic natural secretion patterns.Conclusions: The results demonstrate that when performing studies or experiments these factors can result in alteration of the pain or behavioral response, as well as, joint structure and inflammatory response. Therefore, these factors must be considered when designing experiments focused on determining the mechanisms of action for sex steroids.

Error in Figure/TableIn the original article, there was a mistake in Figure 3B as published. The value of mg should have been µg. The corrected Figure 3B appears below. The authors apologize for this error and state that this does not change the scientific conclusions of the article in any way. The original article has been updated.Text CorrectionIn the original article, there was an error. The value of mg should have been µg.A correction has been made to the Results section, sub-Section Enzyme-linked immunosorbent assay (ELISA).Fresh thalamic tissue punches from all five animals in experiment #3 (Table 1) was stored in liquid nitrogen until analysis. Tissue was placed in 250 µl of T-Per tissue protein extraction reagent containing Halt Protease Inhibitor and ground (Thermo Scientific, Rockford, IL). Ground samples were frozen and thawed, followed by centrifugation and decanting of the supernatant. Quantitation of VGAT in the supernatant was completed on duplicate 100 µl samples of supernatant using an SLC32A1 (VGAT) ELISA following the manufacturer’s directions (Cusabio, catalog #CSB-EL021578MO). Total protein was determined in each sample using a BCA protein assay (Thermo Scientific, Waltham, WA). Values represent the pg of VGAT per µg of total protein.The authors apologize for this error and state that this does not change the scientific conclusions of the article in any way. The original article has been updated.

Optogenetic manipulation is uniquely useful in unraveling the functional organization of neuronal circuits in the central nervous system by enabling reversible gain- or loss-of-function of discrete populations of neurons within restricted brain regions. This state-of-the-art technology can produce circuit-specific neuromodulation by overexpressing light-sensitive proteins (opsins) in particular cell types of interest. Here, we discuss the principle of optogenetic manipulation and its application in pain research using animal models, and we also discuss how to potentially use optogenetic stimulation in the treatment of migraine headache in the future.

Herpes zoster or shingles is the result of varicella zoster virus (VZV) infection and often results in chronic pain that lasts for months after visible symptoms subside. Testosterone often attenuates pain in males. Previous work demonstrates ovarian estrogen effects γ-aminobutyric acid (GABA) signaling in the thalamus, reducing pain but the role of testosterone within the thalamus is currently unknown. Because aromatase affects pain and is present in the thalamus we tested a hypothesis that testosterone converted to estrogen in the thalamus attenuates herpes zoster induced pain. To address this hypothesis, male Sprague-Dawley rats received whisker pad injection of either MeWo cells or MeWo cells containing VZV. To reduce aromatase derived estrogen in these animals we injected aromatase inhibitor letrozole systemically or infused it into the thalamus. To test if estrogen was working through the estrogen receptor (ER) agonist, 4, 4′, 4″-(4-Propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol (PPT) was infused concomitant with letrozole. Motivational and affective pain was measured after letrozole and/or PPT treatment. Vesicular GABA transporter (VGAT) is important in pain signaling. Because estrogen effects VGAT expression we measured its transcript and protein levels after letrozole treatment. Virus injection and letrozole significantly increased the pain response but thalamic infusion of PPT reduced zoster pain. Letrozole increased the number of thalamic neurons staining for phosphorylated ERK (pERK) but decreased VGAT expression. The results suggest in male rats aromatase derived estradiol interacts with the ER to increase VGAT expression and increase neuronal inhibition in the thalamus to attenuate VZV induced pain.

Neuropathic pain represents a challenge to clinicians because it is resistant to commonly prescribed analgesics due to its largely unknown mechanisms. Here, we investigated a descending dopaminergic pathway–mediated modulation of trigeminal neuropathic pain. We performed chronic constriction injury of the infraorbital nerve from the maxillary branch of trigeminal nerve to induce trigeminal neuropathic pain in mice. Our retrograde tracing showed that the descending dopaminergic projection from hypothalamic A11 nucleus to spinal trigeminal nucleus caudalis is bilateral. Optogenetic/chemogenetic manipulation of dopamine receptors D1 and D2 in the spinal trigeminal nucleus caudalis produced opposite effects on the nerve injury–induced trigeminal neuropathic pain. Specific excitation of dopaminergic neurons in the A11 nucleus attenuated the trigeminal neuropathic pain through the activation of D2 receptors in the spinal trigeminal nucleus caudalis. Conversely, specific ablation of the A11 dopaminergic neurons exacerbated such pain. Our results suggest that the descending A11−spinal trigeminal nucleus caudalis dopaminergic projection is critical for the modulation of trigeminal neuropathic pain and could be manipulated to treat such pain.

Varicella zoster virus (VZV) infects the face and can result in chronic, debilitating pain. The mechanism for this pain is unknown and current treatment is often not effective, thus investigations into the pain pathway become vital. Pain itself is multidimensional, consisting of sensory and affective experiences. One of the primary brain substrates for transmitting sensory signals in the face is the ventral posterior medial/posterior lateral thalamus (VPM/VPL). In addition, the anterior cingulate cortex (ACC) has been shown to be vital in the affective experience of pain, so investigating both of these areas in freely behaving animals was completed to address the role of the brain in VZV induced pain. Our lab has developed a place escape avoidance paradigm to measure VZV induced affective pain in the orofacial region of the rat. Using this assay as a measure of the affective pain experience a significant response was observed after VZV injection into the whisker pad and after VZV infusion into the trigeminal ganglion. Local field potentials (LFP) are the summed electrical current from a group of neurons. LFP in both the VPM/VPL and ACC was attenuated in VZV injected rats after inhibition of neuronal activity. This inhibition of VPM/VPL neurons was accomplished using a designer receptor activated by a designer drug (DREADD). Immunostaining showed that cells within the VPM/VPL expressed thalamic glutamatergic vesicle transporter-2, NeuN and DREADD suggesting inhibition occurred primarily in excitable neurons. From these results we conclude 1) that VZV associated pain does not involve a mechanism exclusive to the peripheral nerve terminals and 2) can be controlled, in part, by excitatory neurons within the VPM/VPL that potentially modulate the affective experience by altering activity in the ACC.

Most people are initially infected with varicella zoster virus (VZV) at a young age and this infection results in chickenpox. VZV then becomes latent and reactivates later in life resulting in herpes zoster (HZ) or “shingles”. Often VZV infects neurons of the trigeminal ganglia to cause ocular problems, orofacial disease and occasionally a chronic pain condition termed post-herpetic neuralgia (PHN). To date, no model has been developed to study orofacial pain related to varicella zoster. Importantly, the incidence of zoster associated pain and PHN is known to be higher in women, although reasons for this sex difference remain unclear. Prior to this work, no animal model was available to study these sex-differences. Our goal was to develop an orofacial animal model for zoster associated pain which could be utilized to study the mechanisms contributing to this sex difference. To develop this model VZV was injected into the whisker pad of rats resulting in IE62 protein expression in the trigeminal ganglia; IE62 is an immediate early gene in the VZV replication program. Similar to PHN patients, rats showed retraction of neurites after VZV infection. Treatment of rats with gabapentin, an agent often used to combat PHN, ameliorated the pain response after whisker pad injection. Aversive behavior was significantly greater for up to 7 weeks in VZV injected rats over control inoculated rats. Sex differences were also seen such that ovariectomized and intact female rats given the lower dose of VZV showed a longer affective response than male rats. The phase of the estrous cycle also affected the aversive response suggesting a role for sex steroids in modulating VZV pain. These results suggest that this rat model can be utilized to study the mechanisms of 1) orofacial zoster associated pain and 2) the sex differences underlying zoster associated pain.