A critical evaluation of TRPA1-mediated locomotor behavior in zebrafish as a screening tool for novel anti-nociceptive drug discovery

Abstract

Current medications inadequately treat the symptoms of chronic pain experienced by over 50 million people in the United States, and may come with substantial adverse effects signifying the need to find novel treatments. One novel therapeutic target is the Transient Receptor Potential A1 channel (TRPA1), an ion channel that mediates nociception through calcium influx of sensory neurons. Drug discovery still relies heavily on animal models, including zebrafish, a species in which TRPA1 activation produces hyperlocomotion. Here, we investigated if this hyperlocomotion follows zebrafish TRPA1 pharmacology and evaluated the strengths and limitations of using TRPA1-mediated hyperlocomotion as potential preclinical screening tool for drug discovery. To support face validity of the model, we pharmacologically characterized mouse and zebrafish TRPA1 in transfected HEK293 cells using calcium assays as well as in vivo. TRPA1 agonists and antagonists respectively activated or blocked TRPA1 activity in HEK293 cells, mice, and zebrafish in a dose-dependent manner. However, our results revealed complexities including partial agonist activity of TRPA1 antagonists, bidirectional locomotor activity, receptor desensitization, and off-target effects. We propose that TRPA1-mediated hyperlocomotion in zebrafish larvae has the potential to be used as in vivo screening tool for novel anti-nociceptive drugs but requires careful evaluation of the TRPA1 pharmacology.

Why did the invasive walking catfish cross the road? Terrestrial chemoreception described for the first time in a fish

“…Furthermore, annelid worms are a favourite food item of C. batrachus (Das, 1928) and were found by a survey responder to be in the stomachs of many C. batrachus that were on land and surrounded by annelids. Quinine is a bitter compound that elicits negative taste responses in fish when mixed with food (Lamb & Finger, 1995), and AITC in solution targets TRPA1 nociceptors on solitary chemosensory cells in fish, which are found on the bodies of C. batrachus (Curtright et al, 2015;Hinman et al, 2006;Ko et al, 2019;Kotrschal, 1995), but C. batrachus showed no orientation response to either substance on the substrate. Alternatively, quinine and AITC are barely soluble and insoluble in water, respectively.…”

“…Quinine is a bitter compound that elicits negative taste responses in fish when mixed with food (Lamb & Finger, 1995), and AITC in solution targets TRPA1 nociceptors on solitary chemosensory cells in fish, which are found on the bodies of C. batrachus (Curtright et al ., 2015; Hinman et al ., 2006; Ko et al ., 2019; Kotrschal, 1995), but C. batrachus showed no orientation response to either substance on the substrate. Therefore, C. batrachus may not use these compounds for terrestrial orientation.…”

“…Quinine was chosen because it elicits a negative taste response in fish (Lamb & Finger, 1995). AITC was used to determine the role of nociception in terrestrial orientation because AITC activates TRPA1 nociceptors in fish, causing an aversive response (Curtright et al ., 2015; Hinman et al ., 2006; Ko et al ., 2019). Due to the low solubility of quinine (0.5 g/l) and the insolubility of AITC in water, supersaturated solutions of these substances were created to aversive concentrations, based on the literature (Ko et al ., 2019; Lamb & Finger, 1995), and then shaken in spray bottles immediately before dispersion onto the TA to ensure even coverage.…”

TRPA1 Channel Activation Inhibits Motor Activity in the Mouse Colon

“…It's unlikely that the concentration of HC-030031 in these experiments was too low to have any significant effect upon basal CPMC activity. Whilst HC-030031 is a relatively weak antagonist of murine TRPA1 channels (pA 2 ∼ 5.5; Ko et al, 2019) the antagonist was tested at an upper concentration shown to be specific for blocking TRPA1 (Capasso et al, 2012) and to cause almost complete inhibition of TRPA1 mediated Ca 2+ currents in HEK cells expressing TRPA1 (McNamara et al, 2007).…”

Identification of compounds producing non-visual photosensation via TRPA1 in zebrafish

“…Although previous studies had identified optovin analogues 37 , it was unclear whether TRPA1-mediated nonvisual photosensation also occurred with other, more structurally diverse compounds. Through this study, we have demonstrated that there are multiple novel compounds with structural and photochemical properties that differ greatly from those of optovin but are similarly able to induce motor activity in response to light.…”