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Diffusion-weighted Imaging (DWI) is an effective and state-of-the-art neuroimaging method that non-invasively reveals the microstructure and connectivity of tissues. Recently, novel applications of the DWI technique in studying large brains through ex-vivo imaging enabled researchers to gain insights into the complex neural architecture in different species such as those of Perissodactyla (e.g., horses and rhinos), Artiodactyla (e.g., bovids, swines, and cetaceans), and Carnivora (e.g., felids, canids, and pinnipeds). Classical invivo tract-tracing methods are usually considered unsuitable for ethical and practical reasons, in large animals or protected species. Ex-vivo DWI-based tractography offers the chance to examine the microstructure and connectivity of formalin-fixed tissues with scan times and precision that is not feasible invivo. This paper explores DWI's application to ex-vivo brains of large animals, highlighting the unique insights it offers into the structure of sometimes phylogenetically different neural networks, the connectivity of white matter tracts, and comparative evolutionary adaptations. Here, we also summarize the challenges, concerns, and perspectives of ex-vivo DWI that will shape the future of the field in large brains. [ABSTRACT FROM AUTHOR]

Introduction: The entorhinal cortex has been shown to be involved in high-level cognitive functions in terrestrial mammals. It can be divided into two main areas: the lateral entorhinal area (LEA) and the medial entorhinal area (MEA). Understanding of its structural organization in cetaceans is particularly important given the extensive evidence for their cognitive abilities. The present study describes the cytoarchitectural and immunohistochemical properties of the entorhinal cortex of the bottlenose dolphin (Tursiops truncatus, Montagu, 1821), perhaps the most studied cetacean species and a paradigm for dolphins and other small cetaceans. Methods: Four bottlenose dolphins' entorhinal cortices were processed. To obtain a precise overview of the organization of the entorhinal cortex we used thionin staining to study its laminar and regional organization, and immunoperoxidase technique to investigate the immunohistochemical distribution of three most commonly used calcium-binding proteins (CBPs), calbindin D-28k (CB), calretinin (CR) and parvalbumin (PV). Entorhinal cortex layers thickness were measured, morphological and morphometric analysis for each layer were conducted and statistically compared. Results: Six layers in both the LEA and MEA were identified. The main difference between the LEA and the MEA is observed in layers II and III: the neurons in layer II of the LEA were denser and larger than the neurons in layer II of MEA. In addition, a relatively cell-free zone between layers II and III in LEA, but not in MEA, was observed. The immunohistochemical distribution of the three CBPs, CB, CR and PV were distinct in each layer. The immunostaining pattern of CR, on one side, and CB/PV, on the other side, appeared to be distributed in a complementary manner. PV and CB immunostaining was particularly evident in layers II and III, whereas CR immunoreactive neurons were distributed throughout all layers, especially in layers V and VI. Immunoreactivity was expressed by neurons belonging to different morphological classes: All CBPs were expressed in nonpyramidal neurons, but CB and CR were also found in pyramidal neurons. Discussion: The morphological characteristics of pyramidal and non-pyramidal neurons in the dolphin entorhinal cortex are similar to those described in the entorhinal cortex of other species, including primates and rodents. Interestingly, in primates, rodents, and dolphins, most of the CBP-containing neurons are found in the superficial layers, but the large CR-ir neurons are also abundant in the deep layers. Layers II and III of the entorhinal cortex contain neurons that give rise to the perforant pathway, which conveys most of the cortical information to the hippocampal formation. From the hippocampal formation, reciprocal projections are directed back to the deep layer of the entorhinal cortex, which distributes the information to the neocortex and subcortical area. Our data reveal that in the dolphin entorhinal cortex, the three major CBPs label morphologically heterogeneous groups of neurons that may be involved in the information flow between entorhinal input and output pathways. [ABSTRACT FROM AUTHOR]

Introduction: The auditory system of dolphins and whales allows them to dive in dark waters, hunt for prey well below the limit of solar light absorption, and to communicate with their conspecific. These complex behaviors require specific and sufficient functional circuitry in the neocortex, and vicarious learning capacities. Dolphins are also precocious animals that can hold their breath and swim within minutes after birth. However, diving and hunting behaviors are likely not innate and need to be learned. Our hypothesis is that the organization of the auditory cortex of dolphins grows and mature not only in the early phases of life, but also in adults and aging individuals. These changes may be subtle and involve sub-populations of cells specificall linked to some circuits. Methods: In the primary auditory cortex of 11 bottlenose dolphins belonging to three age groups (calves, adults, and old animals), neuronal cell shapes were analyzed separately and by cortical layer using custom computer vision and multivariate statistical analysis, to determine potential minute morphological differences across these age groups. morphological differences across these age groups. Results: The results show definite changes in interneurons, characterized by round and ellipsoid shapes predominantly located in upper cortical layers. Notably, neonates interneurons exhibited a pattern of being closer together and smaller, developing into a more dispersed and diverse set of shapes in adulthood. Discussion: This trend persisted in older animals, suggesting a continuous development of connections throughout the life of these marine animals. Our findings further support the proposition that thalamic input reach upper layers in cetaceans, at least within a cortical area critical for their survival. Moreover, our results indicate the likelihood of changes in cell populations occurring in adult animals, prompting the need for characterization. [ABSTRACT FROM AUTHOR]

The evolution of the brain in apes and man followed a joint pathway stemming from common ancestors 5–10 million years ago. However, although apparently sharing similar organization and neurochemical properties, association areas of the isocortex remain one of the cornerstones of what sets humans aside from other primates. Brodmann's area 44, the area of Broca, is known for its implication in speech, and thus indirectly is a key mark of human uniqueness. This latero-caudal part of the frontal lobe shows a marked functional asymmetry in humans, and takes part in other complex functions, including learning and imitation, tool use, music and contains the mirror neuron system (MNS). Since the main features in the cytoarchitecture of Broca's area remains relatively constant in hominids, including in our closest relative, the chimpanzee Pan troglodytes , investigations on the finer structure, cellular organization, connectivity and eventual asymmetry of area 44 have a direct bearing on the understanding of the neural mechanisms at the base of our language. The semi-automated image analysis technology that we employed in the current study showed that the structure of the cortical layers of the chimpanzee contains elements of asymmetry that are discussed in relation to the corresponding human areas and the putative resulting disparity of function. [ABSTRACT FROM AUTHOR]