Evaluation of the Functional Roles of TRPA1 Homologs, Painless and dTRPA1, in Chemical Nociception in Drosophila

The detection of harmful chemical irritants is important for the avoidance of potential tissue-damaging and life threatening compounds. There are multiple physiological systems that exist to detect these irritants, and one specific target of these compounds is the TRPA1 channel. Drosophila possess four evolutionary homologs of mammalian TRPA1, two of which are <italic>painless</italic> and <italic>dTRPA1</italic>. These are thought to be involved in chemical nociception, though the specific role of <italic>painless</italic> in the behavioral aversion to the compound, allyl isothiocyanate, is disagreed upon. We have analyzed the behavioral phenotypes of <italic>painless</italic> and <italic>dTRPA1</italic> mutants using the proboscis extension reflex (PER) and two-choice capillary feeding assays, both of which indicated the requirement for each channel in the aversion to AITC. Expression patterns of these two channels were evaluated to determine if there was any overlap in expression between <italic>painless</italic> and <italic>dTRPA1</italic>. We observed a lack of colocalization in the adult CNS. Cell populations were further defined by the identification of cell specific markers. Subsets of <italic>painless</italic>- and <italic>dTRPA1</italic>-expressing cells coexpress the neuropeptides, DH₃₁ and leucokinin, respectively. Additionally, we specifically expressed tetanus toxin, which blocks synaptic transmission, to verify that these drivers were capturing the aversive circuit. However, it is unclear whether <italic>painless</italic> and <italic>dTRPA1</italic> are acting independently or in combination. To assess <italic>painless</italic> and <italic>dTRPA1</italic> cell excitability, the GCaMP transgene was utilized to observe changes in calcium levels. Both <italic>painless</italic>- and <italic>dTRPA1</italic>-expressing cells exhibited significant changes in fluorescence following the application of AITC. Notably, it was determined that activation via AITC occurred in a direct manner following the ectopic expression of <italic>painless</italic> and <italic>dTRPA1</italic> in AKH cells and evaluation of GCaMP responses for <italic>painless</italic> in a <italic>dTRPA1</italic> mutant background. Collectively, these results suggest that each channel is acting independently to detect irritants and that both are required for behavioral aversion.

Grantor dc:publisher

Wake Forest University

Year dc:date.issued

2012

Author dc:creator

• Shoaf, Madison Lee

• Chemical nociception

Language dc:language.iso

en

Handle dc:identifier.uri

http://hdl.handle.net/10339/37290

OAI identifier oai:identifier

oai:wakespace.lib.wfu.edu:10339/37290