Functional organization of visual responses in the octopus optic lobe

Abstract

Cephalopods are highly visual animals with camera-type eyes, large brains, and a rich repertoire of visually guided behaviors. However, the cephalopod brain evolved independently from that of other highly visual species, such as vertebrates, and therefore the neural circuits that process sensory information are profoundly different. It is largely unknown how their powerful but unique visual system functions, since there have been no direct neural measurements of visual responses in the cephalopod brain. In this study, we used two-photon calcium imaging to record visually evoked responses in the primary visual processing center of the octopus central brain, the optic lobe, to determine how basic features of the visual scene are represented and organized. We found spatially localized receptive fields for light (ON) and dark (OFF) stimuli, which were retinotopically organized across the optic lobe, demonstrating a hallmark of visual system organization shared across many species. Examination of these responses revealed transformations of the visual representation across the layers of the optic lobe, including the emergence of the OFF pathway and increased size selectivity. We also identified asymmetries in the spatial processing of ON and OFF stimuli, which suggest unique circuit mechanisms for form processing that may have evolved to suit the specific demands of processing an underwater visual scene. This study provides insight into the neural processing and functional organization of the octopus visual system, highlighting both shared and unique aspects, and lays a foundation for future studies of the neural circuits that mediate visual processing and behavior in cephalopods.

Highlights: The functional organization and visual response properties of the cephalopod visual system are largely unknownUsing calcium imaging, we performed mapping of visual responses in the octopus optic lobeVisual responses demonstrate localized ON and OFF receptive fields with retinotopic organizationON/OFF pathways and size selectivity emerge across layers of the optic lobe and have distinct properties relative to other species.

Figure 1.. Experimental paradigm for calcium imaging of visual responses in the optic lobe

A) Image of a juvenile Octopus bimaculoides. Between their eyes is the central brain complex, including two optic lobes, one behind each eye. An outline of the central brain is shown in burgundy, and the position of the right optic lobe is indicated by the white outline. B) Illustration of octopus visual system anatomy. Bundles of photoreceptor projections exit the back of the eye (left), decussate vertically, and then enter the optic lobe (right) in a retinotopic manner. In the cutaway, the layered structure of the optic lobe can be seen, as it is in our imaging planes. C) Simplified illustration of the anatomy of the layers of the optic lobe. Photoreceptors in the eye send projections through a cell body layer on the surface of the optic lobe (outer granular layer, OGL, purple) to a layer of neuropil below it (plexiform layer, Plex, red). Here most photoreceptors synapse onto projections from cells in the layer below (inner granular layer, IGL, orange). These in turn send projections to the interior of the optic lobe (medulla, Med, yellow). Color code for layers also applies to Figure 1B,D. D) Mean fluorescence image of Cal-520 calcium indicator loading in the optic lobe, demonstrating successful labeling across multiple layers, delineated by dotted lines. Inset shows layers in color overlay. E) Schematic of the experimental set-up. A projector is used to present visual stimuli on the side of the recording chamber, with the preparation underneath the objective of a two-photon microscope. F) Mean timecourse of fluorescence response to a flashed ON spot at one location in the visual field (averaged over five stimulus repetitions), showing spatial organization and temporal dynamics. Stimulus onset and offset are indicated in the gray bar below the frames, and individual frames are shown at 0.2sec intervals. G) Mean fluorescence response across the optic lobe to ON stimuli at three different horizontal locations, averaged across the stimulus duration for five repetitions. H) Mean fluorescence response to an OFF stimulus at the same location as G (middle), averaged across the stimulus duration for five repetitions.

Figure 2.. ON and OFF receptive fields mapped with a sparse noise stimulus

A) Example frames from the sparse noise stimulus used to map receptive fields. Frames were presented consecutively in a randomized order for a 1sec duration each. B) Traces of fluorescence activity at 32 locations across the optic lobe recorded in response to the sparse noise stimulus. C) RFs from four example units, two each for ON (top) and OFF (bottom) components of the stimulus. D) Histogram of RF sizes for ON and OFF stimuli (N=6 experiments). E) Location of units with RFs for ON (red), OFF (blue), or both (magenta) in one session across the optic lobe. F) Fraction of units overall with significant RFs for ON and OFF across the layers of the optic lobe (N=6 experiments).

Figure 3.. Retinotopic organization of visual responses in the octopus optic lobe

A) Example mapping of RFs in the optic lobe of responses to both ON (left) and OFF (right) stimuli. Areas are colored by the position of their RFs along the elevation (top) and azimuth (bottom) as shown by the color scale bars (degrees). B) Scatter plot of RF location for elevation (top) and azimuth (bottom) versus unit location within the optic lobe, for both ON and OFF responses. Adjacent groups of cells responded to adjacent areas of the visual field. C) Mean coefficient of determination for elevation and azimuth maps across all recordings (N=6 experiments). D) Mean scatter in RF location for elevation and azimuth, across all recordings (N=6 experiments). Dashed line shows chance level based on a shuffle control.

Figure 4.. Size selectivity and temporal dynamics across the layers of the optic lobe

A) Mean timecourse of ON (top panels) and OFF (lower panels) responses for each stimulus size, separated by layers of the optic lobe. Response for each layer and luminance are weighted by fraction of units responsive. OGL did not show a significant response to OFF, and therefore was omitted from this figure. Stimulus onset is at t=0 and each frame was presented for 1sec, as shown by gray bars on the x axis (N=6 experiments). B) Mean size tuning curves for ON responses in each layer, normalized to the response to the smallest stimulus (N=6 experiments). C) Mean timecourse of unit responses, averaged across the three sizes of stimulus spots and normalized to the maximum response, for ON (Plex, IGL, Med) and OFF (Med) (N=6 experiments).