Project Grant 2227963

COLLABORATIVE RESEARCH: HIGHER-ORDER PROCESSING IN A PERIPHERAL NEURAL STRUCTURE OF A NUDIBRANCH MOLLUSC -IN NEUROSCIENCE AND IN BIOLOGY MORE BROADLY, UNIVERSAL PRINCIPLES ARE DETERMINED BY COMPARING A WIDE VARIETY OF DIFFERENT ORGANISMS. A TENET OF NEUROSCIENCE IS THAT HIGHER-ORDER COMPUTATIONS ARE PERFORMED BY NEURAL CIRCUITS IN THE CENTRAL NERVOUS SYSTEM (CNS), NOT THE PERIPHERAL NERVOUS SYSTEM (PNS). HOWEVER, THIS MIGHT NOT BE UNIVERSALLY TRUE FOR ALL TYPES OF ANIMALS. THIS PROJECT EXAMINES WHETHER OLFACTORY PROCESSING CIRCUITRY IN A MOLLUSC IS FOUND IN THE PNS RATHER THAN THE CNS, AS IT IS IN OTHER ANIMAL PHYLA SUCH AS VERTEBRATES AND INSECTS. THE PROJECT USES THE NUDIBRANCH MOLLUSC, BERGHIA STEPHANIEAE, WHICH IS A CHARISMATIC LAB-GROWN SPECIES THAT IS AN INEXPENSIVE ALTERNATIVE TO MANY OTHER NEUROSCIENCE LABORATORY SPECIES, THEREBY INCREASING ACCESS TO RESEARCH FOR STUDENTS. BERGHIA, LIKE OTHER GASTROPOD MOLLUSCS HAS LARGE, INDIVIDUALLY IDENTIFIABLE NEURONS IN ITS CNS. HOWEVER, LIKE ITS DISTANT COUSIN, THE OCTOPUS, FAR MORE SMALL NEURONS ARE LOCATED IN ITS PNS, WHICH CONTAINS PERIPHERAL GANGLIA AND SUB-EPITHELIAL NEURONS. ANTHRO?PO?CENTRICISM HAS BIASED RESEARCHERS TO BELIEVE THAT THE BRAIN IS MORE IMPORTANT THAN THE PERIPHERY. HOWEVER, THIS MIGHT NOT BE TRUE FOR ALL TYPES OF NERVOUS SYSTEMS. DETERMINING THE EXTENT TO WHICH PERIPHERAL SYSTEMS IN MOLLUSCS PERFORM COMPUTATIONS THAT WERE THOUGHT TO BE RESERVED FOR THE BRAIN COULD TRANSFORM OUR VIEW OF WHAT ACTUALLY CONSTITUTES A BRAIN. THE GOAL OF THE PROJECT IS TO ELUCIDATE THE NEURAL ARCHITECTURE AND FUNCTION OF THE PERIPHERAL RHINOPHORE COMPLEX, WHICH CONSISTS OF THE RHINOPHORE, A DORSAL CEPHALIC APPENDAGE USED FOR DISTANCE CHEMORECEPTION, AND THE ASSOCIATED RHINOPHORE GANGLION. A CONNECTOMICS AND TRANSCRIPTOMICS APPROACH WILL BE USED TO DETERMINE WHETHER THERE ARE GLOMERULI IN THE RHINOPHORE COMPLEX THAT ARE ORGANIZED LIKE OLFACTORY GLOMERULI IN THE OLFACTORY BULB OF VERTEBRATES AND THE ANTENNAL LOBE OF INSECTS. THERE ARE THREE OBJECTIVES: 1) A CONNECTOME OF THE RHINOPHORE GANGLION WILL BE CONSTRUCTED FROM VOLUME SERIAL ELECTRON MICROSCOPIC IMAGES USING MACHINE-LEARNING ALGORITHMS TO DETERMINE THE NEURAL CIRCUIT MOTIFS THAT IT CONTAINS. CUSTOMIZATION OF MACHINE-LEARNING ALGORITHMS TO THE ULTRASTRUCTURAL FEATURES OF BERGHIA WILL FACILITATE CONNECTOMICS RESEARCH IN OTHER MOLLUSCS. 2) AN ATLAS OF NEURONAL TYPES WILL BE ASSEMBLED, WHICH WILL INCLUDE NEURONS IN THE RHINOPHORE COMPLEX AND NEURONS THAT INPUT TO IT. NEURAL GENE EXPRESSION, DETERMINED FROM SINGLE CELL RNA SEQUENCING, WILL BE MAPPED USING IN SITU HYBRIDIZATION CHAIN REACTION (HCR), IMMUNOHISTOCHEMISTRY (IHC), AND AXON TRACING. A WEB PORTAL WILL PROVIDE ACCESS TO THE ANNOTATED CONNECTOME AND NEURONAL ATLAS, ALLOWING RESEARCHERS AND STUDENTS TO EXPLORE GENES, NEURONS, AND NEURAL CIRCUITRY IN BERGHIA. 3) THE ROLE OF THE RHINOPHORE COMPLEX IN NAVIGATIONAL BEHAVIOR WILL BE EXPLORED WITH BEHAVIORAL ASSAYS AND SELECTIVE LESIONS, TAKING ADVANTAGE OF BERGHIA?S REGENERATIVE ABILITIES. LABORATORY COURSES DEVELOPED USING BERGHIA WILL ALLOW UNDERGRADUATE STUDENTS TO LABEL AND IMAGE NEURONS WITH HCR AND IHC AND PERFORM BEHAVIORAL EXPERIMENTS WITH AUTOMATED TRACKING. THIS WILL ENABLE STUDENTS TO MAKE DISCOVERIES AND TEST HYPOTHESES REGARDING NEURONS AND NEURAL CIRCUITS IN PARALLEL TO THE DISCOVERIES MADE BY LABORATORY RESEARCHERS. THIS AWARD REFLECTS NSF'S STATUTORY MISSION AND HAS BEEN DEEMED WORTHY OF SUPPORT THROUGH EVALUATION USING THE FOUNDATION'S INTELLECTUAL MERIT AND BROADER IMPACTS REVIEW CRITERIA.