Calcium imaging method to visualize the spatial patterns of neural responses in the pygmy squid, Idiosepius paradoxus, central nervous system

Abstract

BackgroundCephalopods exhibit unique behaviors such as camouflage and tactile learning. The brain functions correlated to these behaviors have long been analyzed through behavioral observations of animals subject to surgical manipulation or electrical stimulation of brain lobes. However, physiological methods have rarely been introduced to investigate the functions of each individual lobe, though physiological work on giant axons and slices of the vertical lobe system of the cephalopods have provided deep insights into ion conductance of nerves and long-term synaptic plasticity. The lack of in vivo physiological work is partly due to difficulties in immobilizing the brain which is contained within the soft body and applying calcium indicators to the cephalopod central nervous system. New methodWe here present a calcium imaging method to visualize neural responses in the central nervous system of the smallest squid, Idiosepius paradoxus. ResultsWe injected calcium indicator Cal-520 into the brachial lobes and revealed a spatiotemporal pattern of neural responses to the electrical stimulations of the axial nerve cord in the first arm. Comparison with existing methodsWe established a method to immobilize the central nervous system which is contained within the soft body and record the calcium responses from the intact central nervous system. ConclusionsOur method provides a novel approach to investigate the mechanisms of how the characteristic organization of the cephalopod brain functions to induce their unique behaviors.