Neuron numbers link innovativeness with both absolute and relative brain size in birds.

A longstanding issue in biology is whether the intelligence of animals can be predicted by absolute or relative brain size. However, progress has been hampered by an insufficient understanding of how neuron numbers shape internal brain organization and cognitive performance. On the basis of estimations of neuron numbers for 111 bird species, we show here that the number of neurons in the pallial telencephalon is positively associated with a major expression of intelligence: innovation propensity. The number of pallial neurons, in turn, is greater in brains that are larger in both absolute and relative terms and positively covaries with longer post-hatching development periods. Thus, our analyses show that neuron numbers link cognitive performance to both absolute and relative brain size through developmental adjustments. These findings help unify neuro-anatomical measures at multiple levels, reconciling contradictory views over the biological significance of brain expansion. The results also highlight the value of a life history perspective to advance our understanding of the evolutionary bases of the connections between brain and cognition.

Brain size and neuron numbers drive differences in yawn duration across mammals and birds.

Recent studies indicate that yawning evolved as a brain cooling mechanism. Given that larger brains have greater thermolytic needs and brain temperature is determined in part by heat production from neuronal activity, it was hypothesized that animals with larger brains and more neurons would yawn longer to produce comparable cooling effects. To test this, we performed the largest study on yawning ever conducted, analyzing 1291 yawns from 101 species (55 mammals; 46 birds). Phylogenetically controlled analyses revealed robust positive correlations between yawn duration and (1) brain mass, (2) total neuron number, and (3) cortical/pallial neuron number in both mammals and birds, which cannot be attributed solely to allometric scaling rules. These relationships were similar across clades, though mammals exhibited considerably longer yawns than birds of comparable brain and body mass. These findings provide further evidence suggesting that yawning is a thermoregulatory adaptation that has been conserved across amniote evolution.

EyeCi: Optical clearing and imaging of immunolabeled mouse eyes using light-sheet fluorescence microscopy.

Immunofluorescent imaging is an indispensable technique to study morphology and molecular aspects in tissues. Classical approaches make it necessary to cut physical sections of tissue samples to overcome the limited penetration depth of light, restricting the available information to two dimensions. Recent advances in tissue-clearing techniques enable imaging of fluorescently labeled organs and entire organisms on a cellular level in three dimensions without the need of sectioning. Volume imaging of immunolabeled and cleared tissues started a new era of systems biology, because these techniques provide information on connectivity and circuits, especially in structures with projections in three dimensions such as vascular or nervous systems. The variety of published clearing protocols allows the imaging of every organ with a single exception: the eye. Whole-eye clearing approaches were unsuccessful so far due to the strong pigmentation of the retinal pigment epithelium. Here, we present a new protocol that combines a highly effective melanin bleaching step with solvent-based clearing, termed EyeCi. The protocol is compatible with immunolabeling as demonstrated by the visualization of ocular and retinal vasculature in the intact mouse eye by means of light-sheet fluorescence microscopy. This novel protocol is rapid (1 week) and inexpensive, hence allowing high-throughput, high resolution analysis of vascular architecture of healthy and diseased eyes, in its native, three-dimensional organization within intact eyeballs. Volume imaging of whole cleared eyeballs further enables three-dimensional surface reconstruction and automated quantification of choroidal and retinal vasculature extending ocular imaging to a global level. Thus, EyeCi represents an extension to state-of-the-art light microscopy techniques and is potentially suitable for the investigation of vascular leakage or neovascularization processes.