Cone topography and spectral sensitivity in two potentially trichromatic marsupials, the quokka (Setonix brachyurus) and quenda (Isoodon obesulus)

Abstract

The potential for trichromacy in mammals, thought to be unique to primates, was recently discovered in two Australian marsupials. Whether the presence of three cone types, sensitive to short- (SWS), medium- (MWS) and long- (LWS) wavelengths, occurs across all marsupials remains unknown. Here, we have investigated the presence, distribution and spectral sensitivity of cone types in two further species, the quokka (Setonix brachyurus) and quenda (Isoodon obesulus). Immunohistochemistry revealed that SWS cones in the quokka are concentrated in dorso-temporal retina, while in the quenda, two peaks were identified in naso-ventral and dorso-temporal retina. In both species, MWS/LWS cone spatial distributions matched those of retinal ganglion cells. Microspectrophotometry (MSP) confirmed that MWS and LWS cones are spectrally distinct, with mean wavelengths of maximum absorbance at 502 and 538 nm in the quokka, and at 509 and 551 nm, in the quenda. Although small SWS cone outer segments precluded MSP measurements, molecular analysis identified substitutions at key sites, accounting for a spectral shift from ultraviolet in the quenda to violet in the quokka. The presence of three cone types, along with previous findings in the fat-tailed dunnart and honey possum, suggests that three spectrally distinct cone types are a feature spanning the marsupials.

Figure 1

Immunolabelling of quokka (a,c) and quenda (f,h) retinae. Photomicrographs of wholemounts treated with JH455 (a,f) and JH492 (c,h). Scale bar, 8 μm. Contour maps for each cone type in quokka (b,d) and quenda (g,i): darker shades indicate highest densities, lighter shades, lowest densities. Retinal ganglion cell topographies in quokka (e; modified from Beazley & Dunlop 1983) and quenda (j; modified from Tancred 1981), showing distributions similar to M/LWS cones.

Figure 2

Wholemounted retina of quenda incubated with JH455 and JH492 revealing the presence of unlabelled outer segments (arrow head). Scale bar, 10 μm.

Figure 3

The diprotodont quokka (a) and polyprotodont quenda (a′). Normalized mean averaged absorbance spectra of visual pigments in the quokka (b–d) and quenda (b′–d′), showing pre-bleach spectra (upper traces) with best-fitted visual pigment templates (solid lines) and post-bleach spectra (lower traces) with running averages (solid lines). Scale bar, 10 cm.

Figure 4

Phylogenetic tree of marsupial SWS1 and other opsins generated by the neighbour-joining method (Saitou & Nei 1987) with 1000 bootstrap replications. The calibration bar shows substitutions per site. Amino acid sequences were aligned by Clustal X (Higgins et al. 1996).