Bilateral Whisker Representations in the Primary Somatosensory Cortex in Robo3cKO Mice Are Reflected in the Primary Motor Cortex.

In Robo3cKO mice, midline crossing defects of the trigeminothalamic projections from the trigeminal principal sensory nucleus result in bilateral whisker maps in the somatosensory thalamus and consequently in the face representation area of the primary somatosensory (S1) cortex (Renier et al., 2017; Tsytsarev et al., 2017). We investigated whether this bilateral sensory representation in the whisker-barrel cortex is also reflected in the downstream projections from the S1 to the primary motor (M1) cortex. To label these projections, we injected anterograde viral axonal tracer in S1 cortex. Corticocortical projections from the S1 distribute to similar areas across the ipsilateral hemisphere in control and Robo3cKO mice. Namely, in both genotypes they extend to the M1, premotor/prefrontal cortex (PMPF), secondary somatosensory (S2) cortex. Next, we performed voltage-sensitive dye imaging (VSDi) in the left hemisphere following ipsilateral and contralateral single whisker stimulation. While controls showed only activation in the contralateral whisker barrel cortex and M1 cortex, the Robo3cKO mouse left hemisphere was activated bilaterally in both the barrel cortex and the M1 cortex. We conclude that the midline crossing defect of the trigeminothalamic projections leads to bilateral whisker representations not only in the thalamus and the S1 cortex but also downstream from the S1, in the M1 cortex.

Layers 3 and 4 Neurons of the Bilateral Whisker-Barrel Cortex.

In Robo3R3-5cKO mouse brain, rhombomere 3-derived trigeminal principal nucleus (PrV) neurons project bilaterally to the somatosensory thalamus. As a consequence, whisker-specific neural modules (barreloids and barrels) representing whiskers on both sides of the face develop in the sensory thalamus and the primary somatosensory cortex. We examined the morphological complexity of layer 4 barrel cells, their postsynaptic partners in layer 3, and functional specificity of layer 3 pyramidal cells. Layer 4 spiny stellate cells form much smaller barrels and their dendritic fields are more focalized and less complex compared to controls, while layer 3 pyramidal cells did not show notable differences. Using in vivo 2-photon imaging of a genetically encoded fluorescent [Ca2+] sensor, we visualized neural activity in the normal and Robo3R3-5cKO barrel cortex in response to ipsi- and contralateral single whisker stimulation. Layer 3 neurons in control animals responded only to their contralateral whiskers, while in the mutant cortex layer 3 pyramidal neurons showed both ipsi- and contralateral whisker responses. These results indicate that bilateral whisker map inputs stimulate different but neighboring groups of layer 3 neurons which normally relay contralateral whisker-specific information to other cortical areas.

Methodological aspects of studying the mechanisms of consciousness.

There are at least two approaches to the definition of consciousness. In the first case, certain aspects of consciousness, called qualia, are considered inaccessible for research from a third person and can only be described through subjective experience. This approach is inextricably linked with the so-called "hard problem of consciousness", that is, the question of why consciousness has qualia or how any physical changes in the environment can generate subjective experience. With this approach, some aspects of consciousness, by definition, cannot be explained on the basis of external observations and, therefore, are outside the scope of scientific research. In the second case, a priori constraints do not constrain the field of scientific investigation, and the best explanation of the experience in the first person is included as a possible subject of empirical research. Historically, in the study of cause-and-effect relationships in biology, it was customary to distinguish between proximate causation and ultimate causation existing in biological systems. Immediate causes are based on the immediate influencing factors [1]. Proximate causation has evolutionary explanations. When studying biological systems themselves, such an approach is undoubtedly justified, but it often seems insufficient when studying the interaction of consciousness and the brain [2,3]. Current scientific communities proceed from the assumption that the physical substrate for the generation of consciousness is a neural network that unites various types of neurons located in various brain structures. Many neuroscientists attach a key role in this process to the cortical and thalamocortical neural networks. This question is directly related to experimental and clinical research in the field of disorder of consciousness. Progress in this area of medicine depends on advances in neuroscience in this area and is also a powerful source of empirical information. In this area of consciousness research, a large amount of experimental data has been accumulated, and in this review an attempt was made to generalize and systematize.

In Vivo Voltage-Sensitive Dye Imaging of Subcortical Brain Function.

The whisker system of rodents is an excellent model to study peripherally evoked neural activity in the brain. Discrete neural modules represent each whisker in the somatosensory cortex ("barrels"), thalamus ("barreloids"), and brain stem ("barrelettes"). Stimulation of a single whisker evokes neural activity sequentially in its corresponding barrelette, barreloid, and barrel. Conventional optical imaging of functional activation in the brain is limited to surface structures such as the cerebral cortex. To access subcortical structures and image sensory-evoked neural activity, we designed a needle-based optical system using gradient-index (GRIN) rod lens. We performed voltage-sensitive dye imaging (VSDi) with GRIN rod lens to visualize neural activity evoked in the thalamic barreloids by deflection of whiskers in vivo. We stimulated several whiskers together to determine the sensitivity of our approach in differentiating between different barreloid responses. We also carried out stimulation of different whiskers at different times. Finally, we used muscimol in the barrel cortex to silence the corticothalamic inputs while imaging in the thalamus. Our results show that it is possible to obtain functional maps of the sensory periphery in deep brain structures such as the thalamic barreloids. Our approach can be broadly applicable to functional imaging of other core brain structures.

Thalamic NMDA receptor function is necessary for patterning of the thalamocortical somatosensory map and for sensorimotor behaviors.

NMDARs play a major role in patterning of topographic sensory maps in the brain. Genetic knock-out of the essential subunit of NMDARs in excitatory cortical neurons prevents whisker-specific neural pattern formation in the barrel cortex. To determine the role of NMDARs en route to the cortex, we generated sensory thalamus-specific NR1 (Grin1)-null mice (ThNR1KO). A multipronged approach, using histology, electrophysiology, optical imaging, and behavioral testing revealed that, in these mice, whisker patterns develop in the trigeminal brainstem but do not develop in the somatosensory thalamus. Subsequently, there is no barrel formation in the neocortex yet a partial afferent patterning develops. Whisker stimulation evokes weak cortical activity and presynaptic neurotransmitter release probability is also affected. We found several behavioral deficits in tasks, ranging from sensorimotor to social and cognitive. Collectively, these results show that thalamic NMDARs play a critical role in the patterning of the somatosensory thalamic and cortical maps and their impairment may lead to pronounced behavioral defects.

Sound frequency representation in cat auditory cortex.

Using the intrinsic signal optical recording technique, we reconstructed the two-dimensional pattern of stimulus-evoked neuronal activities in the auditory cortex of anesthetized and paralyzed cats. The average magnitude of intrinsic signal in response to a pure tone stimulus increased steadily as the sound pressure level increased. A detailed analysis demonstrated that the evoked signals at early frames were scaled by the sound pressure level, which in turn indicated the presence of a minimum level of sound pressure beyond which stimulus-related intrinsic signal can be generated. Intrinsic signals evoked significantly by pure tone stimuli of different frequencies were localized and arranged in an orderly manner in the middle ectosylvian gyrus, which indicates that the primary auditory field (AI) is tonotopically organized. The arrangement of optimal frequencies obtained from optical recordings of the same auditory cortex, which were conducted on different days, was highly reproducible. Furthermore, other auditory fields surrounding AI in the recorded area were allocated based on the observed tonotopicity. We also conducted unit recordings on the cats used for optical recording with the same set of acoustic stimuli. The gross feature of the arrangement of optimal frequencies determined by unit recordings agreed with the tonotopic arrangement determined by the optical recording, although the precise agreement was not obtained.

Intrinsic optical signals from rat primary auditory cortex in response to sound stimuli presented to contralateral, ipsilateral and bilateral ears.

In the auditory cortex, primitive features of acoustic stimuli are represented for auditory scene analysis. A typical example of a feature representation is the tonotopic map, in which sound frequencies are spatially arranged in an orderly manner. Some neurons in the auditory cortex are sensitive to sound source location, which is another important feature for auditory scene analysis. In the present study, using the intrinsic optical imaging technique, we attempted to visualize the two-dimensional pattern of neuronal population responses in the primary auditory cortex of rats to pure tones presented at various frequencies and sound intensities. The observed arrangements of sound frequencies were consistent with those obtained by electrophysiological mapping, which indicates that our intrinsic optical recording can visualize populational responses of neurons. We also found different temporal patterns of intrinsic signals elicited in response to contralateral, ipsilateral, and bilateral ear stimulations. Finally we try to explain the observed differential time courses of intrinsic signal responses from the theoretical point of view on the conduction of neural activities, based on the so far anatomically identified neural pathways in the rodent auditory system.