Luiz Fernando Ferrari

Animal models of disease are frequently used to test the safety and efficacy of new treatments and to explore mechanistic insights. The most useful models are those that accurately predict therapeutic success in humans of both pharmacologic and nonpharmacologic interventions. Here, we tested the predictive validity of the Dahl salt-sensitive (SS) rat strain as a model of fibromyalgia syndrome (FMS), a chronic disorder characterized by widespread musculoskeletal pain, fatigue, and mood disorders. Using the Fibromyalgia Analog Model (FAM) index, a multivariate statistical tool that quantifies the magnitude of fibromyalgia traits in an animal, we assessed the effectiveness of both established and investigational pharmacologic and nonpharmacologic therapies in SS rats. FAM scores were higher in female SS rats than males ( P < 0.0001), indicating more robust fibromyalgia-like traits, matching the higher prevalence of FMS in women. When compared to controls, treatments with the FDA-approved drugs for FMS milnacipran and pregabalin (both P < 0.0001, in males and females), as well as the investigational therapies metformin ( P < 0.0001) and green light exposure ( P = 0.0034 and P = 0.0002 for males and females, respectively) effectively decreased the FAM scores, showing reduction of FMS-like symptoms in SS rats. In contrast, indomethacin and the mu-opioid agonist DAMGO displayed limited efficacy ( P = 0.0239 and P = 0.0523, respectively), matching the relative effectiveness of these treatments in humans, justifying why they are not usually prescribed for patients with FMS. The results validate the SS rat as a predictive model in testing pharmacologic and nonpharmacologic therapies for FMS treatment, illustrating the utility of the FAM index in assessing treatment outcomes.

Chronic pain is a multifactorial condition often accompanied by comorbidities such as anxiety, depression, and cardiovascular dysfunction. Traditional injury-based models have provided valuable mechanistic insights but are limited in their ability to capture the spontaneous, polygenic, and systemic nature of human chronic pain. Inherited pain models, such as consomic rat strains, transgenic mice, and recombinant inbred panels, offer a unique advantage towards bridging this translational gap: they enable the study of pain-related mechanisms in the absence of experimental injury, reducing confounding effects and better reflecting clinical complexity. These models serve as powerful platforms to investigate neuroimmune signaling, oxidative stress, and epigenetic regulation, and to explore how these pathways interact with sex, stress, and systemic comorbidities. Importantly, while referred to as “inherited pain models,” these systems are not designed to model pain transmission across generations, but rather to uncover genetically driven susceptibility to pain and its mechanistic basis. Many of the mechanisms identified in these models overlap with findings from human genome-wide association studies, reinforcing their translational relevance. Beyond mechanistic discovery, inherited pain models can be used for the identification of biomarkers, the study of gene-environment interactions, and the development of mechanism-based therapies. Integration with multi-omics technologies and patient-derived systems further enhance their utility. This review highlights how these models are reshaping the field by enabling biologically informed approaches to diagnosis, prevention, and treatment, thus laying the foundations for a more precise and proactive era in pain medicine.

Fibromyalgia syndrome (FMS) is clinically diagnosed using a widespread pain index and symptom severity scores. Despite this, multivariate statistical models using clinical data consistently suggest the presence of single overarching factor that can capture much of the variation across a spectrum of biomarkers for FMS, potentially aiding diagnosis. We hypothesized that a comparable single index could be developed for preclinical FMS rat models to quantitatively compare and rigorously test new candidates for improved translational potential and therapeutic screening. We used male and female reserpine-treated Sprague Dawley (SD) rats to produce the fibromyalgia analog model (FAM) index, a tool to systematically determine the robustness of potential models of FMS. Features associated with FMS were assessed by behavior tests, including widespread nociceptive sensitivity evaluated on the hind paws (“paw”) and gastrocnemius muscles (“mus”) using a Randall–Selitto device, and face (“VF”) using von Frey filaments; depression (by the forced swim test [FST]); anxiety (using the Elevated Zero Maze [EZM]); and fatigue (using a Home Cage Monitoring system [HCM]). A multivariate statistical analysis was used to determine the FAM index and document internal and external model validity. The index equation was calculated to be F = 0.410 × rz _paw + 0.618 × rz _mus + 0.429 × rz _VF + 0.455 × z _FST + 0.183 × RZ_EZM + 0.160 × rz _HCM, where each z is a standardized score. Our data showed the ability of the FAM index to differentiate saline-treated SD rats as a control, “normal” group, vs either acidic saline–injected SD or Dahl S rats as models displaying fibromyalgia-like behaviors.

Nerve growth factor (NGF) exerts multiple functions on target neurons throughout development. The recent discovery of a point mutation leading to a change from arginine to tryptophan at residue 100 in the mature NGFβ sequence (NGF^R100W) in patients with hereditary sensory and autonomic neuropathy type V (HSAN V) made it possible to distinguish the signaling mechanisms that lead to two functionally different outcomes of NGF: trophic versus nociceptive. We performed extensive biochemical, cellular, and live-imaging experiments to examine the binding and signaling properties of NGF^R100W. Our results show that, similar to the wild-type NGF (wtNGF), the naturally occurring NGF^R100Wmutant was capable of binding to and activating the TrkA receptor and its downstream signaling pathways to support neuronal survival and differentiation. However, NGF^R100Wfailed to bind and stimulate the 75 kDa neurotrophic factor receptor (p75^NTR)-mediated signaling cascades (i.e., the RhoA-Cofilin pathway). Intraplantar injection of NGF^R100Winto adult rats induced neither TrkA-mediated thermal nor mechanical acute hyperalgesia, but retained the ability to induce chronic hyperalgesia based on agonism for TrkA signaling. Together, our studies provide evidence that NGF^R100Wretains trophic support capability through TrkA and one aspect of its nociceptive signaling, but fails to engage p75^NTRsignaling pathways. Our findings suggest that wtNGF acts via TrkA to regulate the delayed priming of nociceptive responses. The integration of both TrkA and p75^NTRsignaling thus appears to regulate neuroplastic effects of NGF in peripheral nociception.SIGNIFICANCE STATEMENTIn the present study, we characterized the naturally occurring nerve growth factor NGF^R100Wmutant that is associated with hereditary sensory and autonomic neuropathy type V. We have demonstrated for the first time that NGF^R100Wretains trophic support capability through TrkA, but fails to engage p75^NTRsignaling pathways. Furthermore, after intraplantar injection into adult rats, NGF^R100Winduced neither thermal nor mechanical acute hyperalgesia, but retained the ability to induce chronic hyperalgesia. We have also provided evidence that the integration of both TrkA- and p75^NTR-mediated signaling appears to regulate neuroplastic effects of NGF in peripheral nociception. Our study with NGF^R100Wsuggests that it is possible to uncouple trophic effect from nociceptive function, both induced by wild-type NGF.

Systemic fentanyl induces hyperalgesic priming, long-lasting neuroplasticity in nociceptor function characterized by prolongation of inflammatory mediator hyperalgesia. To evaluate priming at both nociceptor terminals, we studied, in male Sprague Dawley rats, the effect of local administration of agents that reverse type I (protein translation) or type II [combination of Src and mitogen-activated protein kinase (MAPK)] priming. At the central terminal, priming induced by systemic, intradermal, or intrathecal fentanyl was reversed by the combination of Src and MAPK inhibitors, but at the peripheral terminal, it was reversed by the protein translation inhibitor. Mu-opioid receptor (MOR) antisense prevented fentanyl hyperalgesia and priming. To determine whether type I and II priming occur in the same population of neurons, we used isolectin B4–saporin or [Sar⁹, Met(O₂)¹¹]-substance P–saporin to deplete nonpeptidergic or peptidergic nociceptors, respectively. Following intrathecal fentanyl, central terminal priming was prevented by both saporins, whereas that in peripheral terminal was not attenuated even by their combination. However, after intradermal fentanyl, priming in the peripheral terminal requires both peptidergic and nonpeptidergic nociceptors, whereas that in the central terminal is dependent only on peptidergic nociceptors. Pretreatment with dantrolene at either terminal prevented fentanyl-induced priming in both terminals, suggesting communication between central and peripheral terminals mediated by intracellular Ca²⁺signaling.In vitroapplication of fentanyl increased cytoplasmic Ca²⁺concentration in dorsal root ganglion neurons, which was prevented by pretreatment with dantrolene and naloxone. Therefore, acting at MOR in the nociceptor, fentanyl induces hyperalgesia and priming rapidly at both the central (type II) and peripheral (type I) terminal and this is mediated by Ca²⁺signaling.SIGNIFICANCE STATEMENTFentanyl, acting at the μ-opioid receptor (MOR), induces hyperalgesia and hyperalgesic priming at both the central and peripheral terminal of nociceptors and this is mediated by endoplasmic reticulum Ca²⁺signaling. Priming in the central terminal is type II, whereas that in the peripheral terminal is type I. Our findings may provide useful information for the design of drugs with improved therapeutic profiles, selectively disrupting individual MOR signaling pathways, to maintain an adequate long-lasting control of pain.

Repeated stimulation of mu-opioid receptors (MORs), by an MOR-selective agonist DAMGO induces type II priming, a form of nociceptor neuroplasticity, which has 2 components: opioid-induced hyperalgesia (OIH) and prolongation of prostaglandin-E₂(PGE₂)-induced hyperalgesia. We report that intrathecal antisense knockdown of the MOR in nociceptors, prevented theinductionof both components of type II priming. Type II priming was also eliminated by SSP-saporin, which destroys the peptidergic class of nociceptors. Because the epidermal growth factor receptor (EGFR) participates in MOR signaling, we tested its role in type II priming. The EGFR inhibitor, tyrphostin AG 1478, prevented theinductionof prolonged PGE₂-induced hyperalgesia, but not OIH, when tested out to 30 days after DAMGO. However, even when repeatedly injected, an EGFR agonist did not induce hyperalgesia or priming. A phosphopeptide, which blocks the interaction of Src, focal adhesion kinase (FAK), and EGFR, also prevented DAMGO-induced prolongation of PGE₂hyperalgesia, but only partially attenuated theinductionof OIH. Inhibitors of Src and mitogen-activated protein kinase (MAPK) also only attenuated OIH. Inhibitors of matrix metalloproteinase, which cleaves EGF from membrane protein, markedly attenuated theexpression, but did not prevent theinduction, of prolongation of PGE₂hyperalgesia. Thus, although theinductionof prolongation of PGE₂-induced hyperalgesia at the peripheral terminal of peptidergic nociceptor is dependent on Src, FAK, EGFR, and MAPK signaling, Src, FAK, and MAPK signaling is only partially involved in theinductionof OIH.

We studied, in male Sprague Dawley rats, the role of the cognate hyaluronan receptor, CD44 signaling in the antihyperalgesia induced by high molecular weight hyaluronan (HMWH). Low molecular weight hyaluronan (LMWH) acts at both peptidergic and nonpeptidergic nociceptors to induce mechanical hyperalgesia that is prevented by intrathecal oligodeoxynucleotide antisense to CD44 mRNA, which also prevents hyperalgesia induced by a CD44 receptor agonist, A6. Ongoing LMWH and A6 hyperalgesia are reversed by HMWH. HMWH also reverses the hyperalgesia induced by diverse pronociceptive mediators, prostaglandin E₂, epinephrine, TNFα, and interleukin-6, and the neuropathic pain induced by the cancer chemotherapy paclitaxel. Although CD44 antisense has no effect on the hyperalgesia induced by inflammatory mediators or paclitaxel, it eliminates the antihyperalgesic effect of HMWH. HMWH also reverses the hyperalgesia induced by activation of intracellular second messengers, PKA and PKC_ε, indicating that HMWH-induced antihyperalgesia, although dependent on CD44, is mediated by an intracellular signaling pathway rather than as a competitive receptor antagonist. Sensitization of cultured small-diameter DRG neurons by prostaglandin E₂is also prevented and reversed by HMWH. These results demonstrate the central role of CD44 signaling in HMWH-induced antihyperalgesia, and establish it as a therapeutic target against inflammatory and neuropathic pain.SIGNIFICANCE STATEMENTWe demonstrate that hyaluronan (HA) with different molecular weights produces opposing nociceptive effects. While low molecular weight HA increases sensitivity to mechanical stimulation, high molecular weight HA reduces sensitization, attenuating inflammatory and neuropathic hyperalgesia. Both pronociceptive and antinociceptive effects of HA are mediated by activation of signaling pathways downstream CD44, the cognate HA receptor, in nociceptors. These results contribute to our understanding of the role of the extracellular matrix in pain, and indicate CD44 as a potential therapeutic target to alleviate inflammatory and neuropathic pain.