The research presented in this work explores the application of microfluidics to the field of proteomics through the design of a multi-channel microfluidic platform and the investigation of individual components of the system. The design of this microfluidic device allows the integration of several protein sample preparation steps for automated electrospray ionization mass spectrometric (ESI-MS) analysis, including protein separation, fractionation and collection, preconcentration and cleanup, and protein digestion. In order for the multi-channel system to function properly, I first evaluated each individual component of the device. Several areas were explored: (i) optimization of polymer monolith for solid-phase extraction (SPE) preconcentration; (ii) investigation of cationic coatings for microchannel surface modification to facilitate positive electrospray of peptides and proteins for chip-MS coupling; (iii) combination of the hydrophobic monolith and the PolyE-323 coating in a single channel device for on-chip SPE and on-bed tryptic digestion with on-line coupling to ESI-MS. Multiplexed microfluidic devices for protein analysis, which integrate a series of microfluidic features, were then designed, built and tested. The multiplexed microfluidic architecture employed a separation channel, a fractionator, an array of microchambers to accommodate monolithic polymer for SPE preconcentration, and an elution channel for the detection of eluted sample using fluorescence detector or mass spectrometer. The performance of the multiplexed devices for integration of multiple analytical steps was explored with sequential fractionation, collection, and elution of fluorescent sample, evaluating the ability to trap and release individual fractions without cross-contamination. Thorough analysis of each of the individual components on the multiplexed microfluidic platform provides valuable insights into the design of such systems, which brings us closer to our final goal of a proteomic processing microchip.

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Many passerine birds, particularly corvids, are known to express complex cognitive skills comparable to those observed in primates. In order to examine how these similarities are reflected at the cellular level, I counted neurons and nonneuronal cells in passerine brains using the isotropic fractionator method. I show that, in these birds, neuronal numbers scale almost isometrically with telencephalic size, i.e., the average neuron size shows little increase and neuronal density decreases minimally as brains get larger. Neuronal densities in the passerine telencephalon exceed those observed in the primate cerebral cortex by a factor of 3-6. As a result, the number of telencephalic neurons in the Common Raven (Corvus corax) equals those observed in the cerebral cortex of small monkeys. The cerebellum features similar scaling rules. However, because the relative size of the cerebellum is smaller than in mammalian brains, cerebellar neurons make a much smaller proportion of total brain neurons than in mammals. In contrast to the little variation in neuronal densities in telencephalon and cerebellum, the density of neurons rapidly decreases with increasing structure size in the diencephalon, optic tectum and brain stem. For all examined brain structures, the densities of nonneuronal cells remain constant...

Galliform birds (Galliformes) make up together with anseriform birds (Anseriformes) the clade Galloanserae, the sister group of Neoaves and the most basal clade of Neognathae. However, to date no quantitative data on cellular composition of their brains have been available. Here, I used the isotropic fractionator to determine numbers of neurons and non-neuronal cells in specific brain regions of 15 species of galliform birds. I find that cellular scaling rules for galliforms differ starkly from those for songbirds and parrots. When compared to these crown avian lineages, galliform birds feature lower degree of encephalization, a proportionally smaller telencephalon, small telencephalic and dominant cerebellar neuronal fractions, generally lower neuronal densities and larger glia/neuron ratios. Consequently, their brains and especially their forebrains harbor much smaller absolute numbers of neurons than those of equivalently sized songbird and parrots, the fact that undoubtedly constrains cognitive abilities of galliforms. However, this not to say that galliform birds are "bird brains" with low numbers of neurons and a limited ability to learn. Because they have high neuronal densities, their relatively small brains contain about equal numbers of neurons as brains of equivalently sized rodents and...

The social brain hypothesis (SBH) posits that complex social environments exert a major selection pressure driving the evolution of large brains and intelligence. The hypothesis was first proposed to explain the remarkable cognitive abilities of primates and has since been extended to other vertebrate groups and gained a substantial popularity. Nevertheless, the empirical support is equivocal in virtually every group where the hypothesis has been tested. In this thesis, the SBH is tested in the African mole-rats (Bathyergidae). Mole-rats share a subterranean mode of life and similar ecologies while covering the whole social spectrum, from solitary to "eusocial". The number of brain neurons is considered a better proxy for intelligence than relative or absolute brain size. Therefore, a novel approach, the isotropic fractionator, was used to estimate the total number of neurons and other cells in five brain parts (olfactory bulbs, cerebral cortex, cerebellum, diencephalon and basal ganglia, brain stem) of eleven bathyergid species. This simultaneously allows for examining if and how mole-rats differ from other rodents with respect to brain cellular scaling rules. We found that, contrary to expectations, mole-rats generally conform to these rules, with a few exceptions. They tend to have higher...

Recent comparative studies have shown that bird brains, although small, have a high processing capacity. The brains of parrots and songbirds have higher neuronal densities than brains of mammals; especially large parrots and corvids compete with or even outnumber primates by the number of telencephalic neurons. However, the processing capacity of the avian brain appears to differ significantly between various phylogenetic lineages. Basal groups such as galliform birds have much lower absolute numbers of neurons and lower neuronal densities than songbirds and parrots. In this Master thesis, I used the isotropic fractionator to determine numbers of neurons and non-neural cells in specific brain regions in 19 species of hornbills (Bucerotiformes), woodpeckers (Piciformes) and coraciiform birds (Coraciiformes). The brains of hornbills and woodpeckers (but not coraciiform birds) have numbers of neurons comparable to that of songbirds and parrots and significantly more neurons than equivalently sized brains of pigeons (Columbiformes) and galliform birds (Galliformes). In the crown groups, we can observe similar trends such as a higher degree of encephalization, a proportionally larger telencephalon and increasing percentage of telencephalic neurons. On the contrary, in pigeons and galliform birds, we can...