The wealth of biodiversity in the world’s library of venoms and their toxins represents an enormous untapped resource that could contain the scaffolds for novel therapeutic drugs. The red lionfish (Pterois volitans) is a venomous species of fish originating from the Indo-Pacific but now invasive in many regions, where it poses a significant stress on marine ecosystems and produces one of the most painful stings in the ocean. In a study I completed prior to this thesis, we demonstrated the qualities of the pain elicited by the venom (in mice) as well as its high specificity for its cellular target - nonpeptidergic nociceptors. These cells are responsible for detecting pain in the peripheral nervous system and the venom somehow specifically activates them over other sensory neurons responsible for touch or proprioception. How can the venom target these cells specifically? This was the broad question that led to this doctoral project. The objective of this thesis was to gain insights into the human pain experience of lionfish stings, the proteinaceous toxin components of the venom and its molecular mechanism of action.In chapter 2, I co-designed a detailed pain questionnaire which was completed by over 500 people who had been stung by lionfish to understand the pain they experienced and its impact on their daily lives. This was the first broad-scale study of lionfish stings ever performed and provided key insights into the average duration of pain caused by lionfish stings, causes of stings, locations of stings and other important variables.In chapter 3, I examined the toxin composition of the venom using a combinatorial transcriptomic and proteomic approach. We performed de novo RNA sequencing of the lionfish’s venomous spines and assembled a transcriptome, which we used in mass spectrometry experiments to identify the proteins expressed in the lionfish venom. From there, I characterized the most abundant proteins and transcripts, synthesized toxins and tested them individually for their ability to activate nociceptors. I identified a highly abundant pain-causing toxin, whose sequence is a close match to Apolipoprotein-E , a molecule that has been intrinsically linked to pain sensation but whose function and mechanism of action remains poorly understood. In chapter 4, I examined the mechanism through which the venom activates sensory neurons to understand what receptor is responsible for the venom’s activation of nonpeptidergic nociceptors. I showed that the venom activates nociceptors through activation of P2X3, and triggers ATP release from these cells in order to further activate neighbouring neurons and non-neuronal cells. Antagonism of the P2X3 receptor significantly reduces the pain caused by lionfish venom in mice, suggesting that the venom relies heavily on direct or indirect activation of P2X3 to cause pain. The findings revealed in my thesis form tMcGill University, Montreal, Quebec he basis of a new understanding of the components of lionfish venom, their mechanism of action and the human experience of lionfish stings. It has brought to light potential molecules that may be used in the study of the P2X3 receptor channel, and sheds light on how the lionfish’s venom has evolved to specifically cause pain in its victims