Nociception refers to the ability to detect pain or injury-inducing stimuli, and is critical for an animal?s survival. How painful stimuli are transmitted to the brain can be changed by the brain itself. For example, damaged regions of a body become more sensitive to subsequent stimulation as a way to protect the body from further damage. Alternatively, gently rubbing a painful area of the body can temporarily reduce pain. The goal of this project is to understand the physiological processes that modulate the ways in which painful stimuli are transmitted to the brain, focusing on the effects of endocannabinoids, a class of neurotransmitters known to attenuate pain. In addition, the capacity for certain patterns of neural activity to stimulate the release of endocannabinoid transmitters will be examined. This project will include behavioral experiments, molecular genetic procedures and electrophysiological recordings of synaptic transmission by pain-sensing neurons (nociceptors). It is expected that (1) endocannabinoid transmitters will be found to decrease nociceptive synaptic signaling, which will lead to a decrease in pain-elicited behaviors and (2) that activation of non-pain, touch-sensitive neurons will stimulate the release of endocannabinoid transmitters resulting in a decrease in the effects of painful stimuli. The experiments in this project are critical to developing an understanding of a fundamental process in neurobiology, how the brain detects pain and how pain signaling can be altered by modulatory processes within the brain itself. This proposal also provides unique training opportunities for graduate and undergraduate students in behavioral, molecular genetics and electrophysiological recording techniques. Furthermore, this project will take place in a state (South Dakota) that is under-represented in terms of federally funded scientific research and at an institution (University of South Dakota) that serves a number of rural and/or first generation college students.
The first goal of this project was to examine the effects of neurotransmitters called endocannabinoids on pain signaling at the level of synaptic transmission. The second goal was to examine the role of a specific type of receptor, called a Transient Receptor Potential Vanilloid or TRPV, in mediating the effects of endocannabinoids. Endocannabinoids are lipid neurotransmitters that typically bind to cannabinoid receptors (the same receptors that the active ingredients in marijuana, THC, bind to). However, endocannabinoids are also known to bind to TRPV channels, which are primarily known for detecting noxious thermal and chemical stimuli (capsaicin is a well-known activator of TRPV channels). These experiments were carried out in the medicinal leech, in which it is possible to carry out detailed recordings of pain vs. non-pain neuro-signaling pathways that cannot easily be replicated in the vertebrate nervous system. An additional advantage of the leech is that it possesses TRPV receptors, but does not possess cannabinoid receptors, making it possible to selectively focus on only the TRPV-mediated component of endocannabinoid modulation without having to resort to pharmacological or genetic manipulations. In fact, it appears that all invertebrates lack cannabinoid receptors, suggesting that TRPV is, in evolutionary terms, the original receptor for these endocannabinoid transmitters. Our findings were the following. Endocannabinoids depressed synaptic transmission by sensory cells that detect painful stimuli. Endocannabinoids also depressed behavioral responses elicited by selective activation of these pain-sensing neurons, indicating that the synaptic effects were relevant at the behavioral level. Low frequency stimulation of non-pain sensory neurons (touch cells) reduced synaptic transmission by pain sensing neurons and behavior elicited by these pain-sensing neurons. This synaptic and behavioral depression was mediated by endocannabinoid activation of TRPV. These findings demonstrate that endocannabinoids and TRPV are part of an endogenous modulatory system for controlling pain signaling in the central nervous system. Surprisingly, endocannabinoids potentiate synaptic transmission by non-pain, pressure-sensitive neurons. These findings suggest that endocannabinoids can have both pain-enhancing and pain-reducing effects. Using pharmacological, behavioral and electrophysiological approaches, we have evidence that the leech does possess TRPV channels in both the periphery, where it is involved in detection of noxious stimuli, and in the central nervous system, where it appears to have a neuromodulatory role. We also have evidence of a related protein, TRPA, in both the periphery and central nervous system. The role of peripheral TRPA is probably sensing noxious stimuli (like TRPV). The role of central TRPA is unknown at this time. To date, this research has produced nine presentations at professional meetings and four peer-reviewed publications with another two articles either in press or under revision, plus an three additional manuscripts in preparation. In terms of broader impacts, this project supported the research of four graduate students and eight undergraduate students. Four of the undergraduate trainees were first generation college students and one was Native American. All of the student trainees had opportunities to present their research either at IdeaFest (the University of South Dakota’s student research symposium), the Society for Neuroscience Annual Meeting, or the Faculty for Undergraduate Neuroscience Symposium. In addition, the principal investigator has presented this work as part of a local TEDx talk (www.youtube.com/watch?v=b32tZfte_JA).
