Analgesia is particularly susceptible to placebo responses. Recent studies in humans have provided important insights into the neurobiology underlying placebo-induced analgesia. However, human studies provide incomplete mechanistic explanations of placebo analgesia because of limited capacity to use cellular, molecular, and genetic manipulations. To address this shortcoming, we describe here the development of a rat model of conditioned analgesia in an operant pain assay. Specifically, rats were conditioned to associate a placebo manipulation with the analgesic effect of 1 mg/kg morphine (s.c.) on facial thermal pain. We found that conditioned (placebo) responding bore three of the hallmarks of placebo-induced analgesia: (1) strong inter-animal variability in the response, (2) suppression by the opiate antagonist naloxone (5 mg/kg, s.c.), and (3) a positive predictive relationship between the unconditioned analgesic effect and the conditioned (placebo) effect. Due to the operant nature of the assay and the use of only a mild noxious thermal stimulus, we suggest these results provide evidence of placebo-induced analgesia in a preclinical model that utilizes an affective behavioral endpoint. This finding may provide opportunities for invasive preclinical studies allowing greater understanding of placebo-induced analgesia, thus paving the way for avenues to harness its benefits.

Clinically many orofacial pain complaints including trigeminal neuralgia, tooth extractions, and pain associated with orthodontic tooth movement involve a mechanical component (i.e., touch sensitivity or pain with injury). Although the characteristics of clinical orofacial pain are well described, evaluation of orofacial pain in animals is challenging. Previously, assessments of trigeminal nerve-mediated nociceptive responses have been limited to methods that assess processing within the brain stem [5,10,17,23], utilizing unlearned behaviors that were elicited by mechanical sensitivity using von Frey filaments [23] or thermal stimulation [9]. Our lab has previously reported the development of a novel thermal operant facial testing system that provides an investigator-independent behavioral assessment system ( Fig 1A). We use operant conflict paradigms to establish a behavioral outcome in which an animal can decide between receiving a reward and escaping an aversive stimulus [13,18]. The current report utilizes a modification of our original operant test system by adding a simple mechanical component. This new modification provides investigators with an opportunity to assess both mechanical and thermal assessments of facial allodynia, hyperalgesia, and pain using the same operant system that we developed previously [15]. This novel mechanical behavioral assessment strategy of orofacial pain could further provide a key step for the advancement of translational pain research, as we now demonstrate the ability to directly assess and compare mechanical versus thermal pain using the same outcomes.The objective of this study was to characterize behavioral responses to facial mechanical stimulation and to see how these responses change under conditions of inflammation and analgesia. Since we are using an adaptation of our established thermal facial operant testing paradigm, we also compared baseline thermal stimuli data (unpublished) collected over the past 6 months using our standard thermal operant system, as described in detail previously [15]. In this current study, we modified standard nickel titanium (NiTi) wires similar to those used in orthodontics for providing tooth movement. The advantage of nickel titanium wires is they have tremendous memory and strength, thus allowing for standardized repeated use of the same wires. Additionally, these wires can conduct electricity, so we were able to modify our existing cage to detect and record actual facial contacts, which is one of the keys * John K. Neubert, D.D.S., Ph.D., University of Florida, 1395 Center Drive, College of Dentistry, Dept. of Orthodontics, Gainesville, FL, jneubert@dental.ufl.edu. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production pro...

We present an operant system for the detection of pain in awake, conscious rodents. The Orofacial Pain Assessment Device (OPAD) assesses pain behaviors in a more clinically relevant way by not relying on reflex-based measures of nociception. Food fasted, hairless (or shaved) rodents are placed into a Plexiglas chamber which has two Peltier-based thermodes that can be programmed to any temperature between 7°C and 60 °C. The rodent is trained to make contact with these in order to access a reward bottle. During a session, a number of behavioral pain outcomes are automatically recorded and saved. These measures include the number of reward bottle activations (licks) and facial contact stimuli (face contacts), but custom measures like the lick/face ratio (total number of licks per session/total number of contacts) can also be created. The stimulus temperature can be set to a single temperature or multiple temperatures within a session. The OPAD is a high-throughput, easy to use operant assay which will lead to better translation of pain research in the future as it includes cortical input instead of relying on spinal reflex-based nociceptive assays.

Recent studies have shown that infraorbital nerve constriction (IoNC)-induced mechanical allodynia has been attenuated by administration of highly purified 150-kDa Botulinum neurotoxin type A (BoNT/A). Here, we extend these studies to determine whether BoNT/A could attenuate IoNC-induced symptoms of thermal hyperalgesia. Instead of testing head withdrawal thresholds, a thermal operant assay was used to evaluate cortical processing of sensory input following IoNC. In this assay, a fasted rat's desire to obtain a food reward (sweetened condensed milk) is coupled to its ability to tolerate facial contact with a warm (45 °C) thermode. Bilateral IoNC decreased the ratio of thermode contact duration/event, which is an indicative of thermal hyperalgesia. BoNT/A injection intradermally in the area of infraorbital nerve (IoN) innervation 7 days after IoNC resulted in decreased number of facial contacts and increased the ratio of contact duration/event (measured at 14 days after IoNC). The BoNT/A (2-200 pg) effects were dose dependent and statistically significant at 100 and 200 pg (P < 0·05). Complete reversal of thermal hyperalgesia symptoms was obtained with a 200-pg dose, without affecting sham rat behaviour. Off-site (neck) injection of BoNT/A did not relieve thermal hyperalgesia, while co-injection of BoNT/A with a neutralising antibody in the area of IoN innervation prevented relief of thermal hyperalgesia. Neither IoNC nor BoNT/A injection affected operant assay parameters with a 24 °C thermode, indicating selectivity of thermal hyperalgesia measurements. These results strongly suggest that intradermal injection of BoNT/A in the area of IoN innervation alleviates IoNC-induced thermal hyperalgesia in an operant assay.

Checkpoint inhibition (CPI) therapies have been proven to be powerful clinical tools in treating cancers. FDA approvals and ongoing clinical development of checkpoint inhibitors for treatment of various cancers highlight the immense potential of checkpoint inhibitors as anti-cancer therapeutics. The occurrence of immune-related adverse events, however, is a major hindrance to the efficacy and use of checkpoint inhibitors as systemic therapies in a wide range of patients. Hence, methods of sustained and tumor-targeted delivery of checkpoint inhibitors are likely to improve efficacy while also decreasing toxic side effects. In this review, we summarize the findings of the studies that evaluated methods of tumor-targeted delivery of checkpoint inhibitors, review their strengths and weaknesses, and discuss the outlook for therapeutic use of these delivery methods.