The legend goes that an Indian who was lost in an Andean forest and was down with fever drank from a pool of stagnant water and found that it tasted bitter. He realized the water must have been contaminated by the neighboring quina-quina tree. He thought he was poisoned, but to his surprise, his fever abated. Ever since, extracts of the bitter bark of the quina-quina tree or quinine have been used as a remedy for fever.
The aversion to bitter stimuli is shared between vertebrates and invertebrates albeit through different mechanisms. In vertebrates, electrical synapses are composed of connexins or gap junctions which are evolutionarily unrelated to the invertebrate innexins, or INXs. The separate evolution of electrical synapses suggests the functional necessity of these channels. For many animals, the ability to sense bitter taste can protect them from ingesting harmful foods. Researchers in the Bai lab (Basic Sciences Division) seek to understand bitter taste sensing in the worm C. elegans, a model invertebrate. In their recent publication in PLoS Genetics, they demonstrate how electrical synapse proteins INX-18 and INX-19 modulate aversive behavior in C. elegans.
Previous research found that cGMP, a small signaling molecule, functions within the sensory neuron known as ASH but was produced in the neighboring neuron known as ASK. In ASH, cGMP was responsible for dampening nociceptive sensitivity, making the animals less sensitive to pain. “We wanted to examine the possibility that INX-18/INX-19 electrical synapses mediate cGMP exchange between ASH and ASK neurons,” said Dr. Jihong Bai.
Lisa Voelker, a graduate student in the lab led that study. Voelker and colleagues generated mutant worms lacking the electrical synapse proteins INX-18 and INX-19. They observed that mutant worms become hypersensitive to the bitter tasting quinine. By complementing the mutant worms with transgenic INX-18/INX-19 in genetic rescue experiments, they were able to elucidate how INX-18/INX-19 functions within the sensory ASH neurons and the neighboring ASK neurons. Specifically, they found that INX19 operates in both ASH and ASK neurons while INX-18 is only required in ASK neurons to maintain quinine sensitivity.
Dr. Bai explained the significance of their findings: “This study shows that electrical synapses modulate sensory responses by passing small signaling molecules such as cGMP and Ca2+. Specifically, we found that the electrical synapse proteins INX-18 and INX-19 function within two sensory neurons (ASK and ASH) to regulate the quinine avoidance response of C. elegans.”
In another series of experiments, the researchers used live-cell imaging of fluorescently-labelled INX-18/19 proteins and examined their localization within the ASH/ASK neurons in vivo. They found that INX-19 in ASK and ASH localizes to the same regions in the nerve ring, suggesting that both sides of ASK-ASH electrical synapses contain INX-19. They also found that INX-18 rarely colocalizes with INX-19 within ASK neurons suggesting that INX-18 and INX-19 do not function in the same synapse. Notably, deletion of INX-18 disrupted the localization of INX-19 while removing INX-19 does not alter INX-18 localization. “These innexins form electrical synapses between ASK and ASH, where INX-19 is a major player, though INX-18 is important for correct localization of INX-19 to ASK synapses,” added Dr. Bai.
Since cGMP signaling is an integral component of the quinine-sensing pathway, the question remained whether INX-18/19 played a role in cGMP exchange between ASH and ASK neurons. They used a fluorescent cGMP reporter and examined quinine-induced cGMP signaling in mutant worms. They found both INX-18 and INX-19 are required for proper cGMP signaling in ASK and ASH neurons.
In summary, the study elucidates how INX-18/19 electrical synapses mediate cGMP exchange between neurons as mechanism underlying a behavioral process allowing C. elegans to taste and avoid bitterness. The study opens the horizon for more questions such as whether “signaling molecules traveling through electrical synapses have the potential to alter behavior outputs from neighboring circuits,” said Dr. Bai.
Voelker L, Upadhyaya B, Ferkey DM, Woldemariam S, L'Etoile ND, Rabinowitch I, Bai J. 2019. INX-18 and INX-19 play distinct roles in electrical synapses that modulate aversive behavior in Caenorhabditis elegans. PLoS Genetics.
Cancer Consortium member Jihong Bai contributed to this research.
This research was funded by the National Institute on Deafness and Other Communication Disorders.