Neurophotonic methods in approach to in vivo animal epileptic models: Advantages and limitations.

Neurophotonic technology is a rapidly growing group of techniques that are based on the interactions of light with natural or genetically modified cells of the neural system. New optical technologies make it possible to considerably extend the tools of neurophysiological research, from the visualization of functional activity changes to control of brain tissue excitability. This opens new perspectives for studying the mechanisms underlying the development of human neurological diseases. Epilepsy is one of the most common brain disorders; it is characterized by recurrent seizures and affects >1% of the world's population. However, how seizures occur, spread, and terminate in a healthy brain is still unclear. Therefore, it is extremely important to develop appropriate models to accurately explore the causal relationship of epileptic activity. The use of neurophotonic technologies in epilepsy research falls into two broad categories: the visualization of neural epileptic activity, and the direct optical influence on neurons to induce or suppress epileptic activity. An optogenetic variant of the classical kindling model of epileptic seizures, in which activatable cells are genetically defined, is called optokindling. Research is also underway concerning the application of neurophotonic techniques for suppressing epileptic activity, aiming to bring these methods into clinical practice. This review aims to systematize and describe new approaches that use combinations of different neurophotonic methods to work with in vivo models of epilepsy. These approaches overcome many of the shortcomings associated with classical animal models of epilepsy and thus increase the effectiveness of developing new diagnostic methods and antiepileptic therapy.

The Anti-Epileptic Effects of Carbenoxolone In Vitro and In Vivo.

Gap junctions (GJs) are intercellular junctions that allow the direct transfer of ions and small molecules between neighboring cells, and GJs between astrocytes play an important role in the development of various pathologies of the brain, including regulation of the pathological neuronal synchronization underlying epileptic seizures. Recently, we found that a pathological change is observed in astrocytes during the ictal and interictal phases of 4-aminopyridin (4-AP)-elicited epileptic activity in vitro, which was correlated with neuronal synchronization and extracellular epileptic electrical activity. This finding raises the question: Does this signal depend on GJs between astrocytes? In this study we investigated the effect of the GJ blocker, carbenoxolone (CBX), on epileptic activity in vitro and in vivo. Based on the results obtained, we came to the conclusion that the astrocytic syncytium formed by GJ-associated astrocytes, which is responsible for the regulation of potassium, affects the formation of epileptic activity in astrocytes in vitro and epileptic seizure onset. This effect is probably an important, but not the only, mechanism by which CBX suppresses epileptic activity. It is likely that the mechanisms of selective inhibition of GJs between astrocytes will show important translational benefits in anti-epileptic therapies.

Behavioral Consequences of a Bifacial Map in the Mouse Somatosensory Cortex.

The whisker system is an important sensory organ with extensive neural representations in the brain of the mouse. Patterned neural modules (barrelettes) in the ipsilateral principal sensory nucleus of the trigeminal nerve (PrV) correspond to the whiskers. Axons of the PrV barrelette neurons cross the midline and confer the whisker-related patterning to the contralateral ventroposteromedial nucleus of the thalamus, and subsequently to the cortex. In this way, specific neural modules called barreloids and barrels in the contralateral thalamus and cortex represent each whisker. Partial midline crossing of the PrV axons, in a conditional Robo3 mutant (Robo3R3-5cKO) mouse line, leads to the formation of bilateral whisker maps in the ventroposteromedial, as well as the barrel cortex. We used voltage-sensitive dye optical imaging and somatosensory and motor behavioral tests to characterize the consequences of bifacial maps in the thalamocortical system. Voltage-sensitive dye optical imaging verified functional, bilateral whisker representation in the barrel cortex and activation of distinct cortical loci following ipsilateral and contralateral stimulation of the specific whiskers. The mutant animals were comparable with the control animals in sensorimotor tests. However, they showed noticeable deficits in all of the whisker-dependent or -related tests, including Y-maze exploration, horizontal surface approach, bridge crossing, gap crossing, texture discrimination, floating in water, and whisking laterality. Our results indicate that bifacial maps along the thalamocortical system do not offer a functional advantage. Instead, they lead to impairments, possibly due to the smaller size of the whisker-related modules and interference between the ipsilateral and contralateral whisker representations in the same thalamus and cortex.SIGNIFICANCE STATEMENT The whisker sensory system plays a quintessentially important role in exploratory behavior of mice and other nocturnal rodents. Here, we studied a novel mutant mouse line, in which the projections from the brainstem to the thalamus are disrupted. This led to formation of bilateral whisker maps in both the thalamus and the cortex. The two whisker maps crowd in a space normally devoted to the contralateral map alone and in a nonoverlapping fashion. Stimulation of the whiskers on either side activates the corresponding region of the map. Mice with bilateral whisker maps perform well in general sensorimotor tasks but show poor performance in specific tests that require whisker-dependent tactile discrimination. These observations indicate that contralateral, instead of bilateral, representation of the sensory space plays a critical role in acuity and fine discrimination during somesthesis.

Nanoparticle-based fluoroionophore for analysis of potassium ion dynamics in 3D tissue models and in vivo.

The imaging of real-time fluxes of K+ ions in live cell with high dynamic range (5-150 mM) is of paramount importance for neuroscience and physiology of the gastrointestinal tract, kidney and other tissues. In particular, the research on high-performance deep-red fluorescent nanoparticle-based biosensors is highly anticipated. We found that BODIPY-based FI3 K+-sensitive fluoroionophore encapsulated in cationic polymer RL100 nanoparticles displays unusually strong efficiency in staining of broad spectrum of cell models, such as primary neurons and intestinal organoids. Using comparison of brightness, photostability and fluorescence lifetime imaging microscopy (FLIM) we confirmed that FI3 nanoparticles display distinctively superior intracellular staining compared to the free dye. We evaluated FI3 nanoparticles in real-time live cell imaging and found that it is highly useful for monitoring intra- and extracellular K+ dynamics in cultured neurons. Proof-of-concept in vivo brain imaging confirmed applicability of the biosensor for visualization of epileptic seizures. Collectively, this data makes fluoroionophore FI3 a versatile cross-platform fluorescent biosensor, broadly compatible with diverse experimental models and that crown ether-based polymer nanoparticles can provide a new venue for design of efficient fluorescent probes.

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.

Recent progress in voltage-sensitive dye imaging for neuroscience.

Voltage-sensitive dye imaging (VSDi) enables visualization of information processing in different areas of the brain with reasonable spatial and temporal resolution. VSDi employs different chemical compounds to transduce neural activity directly into the changes in intrinsic optical signal. Physically, voltage-sensitive dyes (VSDs) are chemical probes that reside in the neural membrane and change their fluorescence or absorbance in response to membrane potential changes. Based on these features, VSDs can be divided into two groups-absorbance and fluorescence. The spatial and temporal resolution of the VSDi is limited mainly by the technical characteristics of the optical imaging setup (e.g., computer and light-sensitive device-charge-coupled device (CCD) camera or photodiode array). In this article, we briefly review the development of the VSD, technique of VSDi and applications in functional brain imaging.

Beyond life: Exploring hemodynamic patterns in postmortem mice brains.

We utilize Laser Speckle Contrast Imaging (LSCI) for visualizing cerebral blood flow in mice during and post-cardiac arrest. Analyzing LSCI images, we noted temporal blood flow variations across the brain surface for hours postmortem. Fast Fourier Transform (FFT) analysis depicted blood flow and microcirculation decay post-death. Continuous Wavelet Transform (CWT) identified potential cerebral hemodynamic synchronization patterns. Additionally, non-negative matrix factorization (NMF) with four components segmented LSCI images, revealing structural subcomponent alterations over time. This integrated approach of LSCI, FFT, CWT, and NMF offers a comprehensive tool for studying cerebral blood flow dynamics, metaphorically capturing the 'end of the tunnel' experience. Results showed primary postmortem hemodynamic activity in the olfactory bulbs, followed by blood microflow relocations between somatosensory and visual cortical regions via the superior sagittal sinus. This method opens new avenues for exploring these phenomena, potentially linking neuroscientific insights with mysteries surrounding consciousness and perception at life's end.

Role of Glial Cells in Neuronal Function, Mood Disorders, and Drug Addiction.

Mood disorders and substance use disorder (SUD) are of immense medical and social concern. Although significant progress on neuronal involvement in mood and reward circuitries has been achieved, it is only relatively recently that the role of glia in these disorders has attracted attention. Detailed understanding of the glial functions in these devastating diseases could offer novel interventions. Here, following a brief review of circuitries involved in mood regulation and reward perception, the specific contributions of neurotrophic factors, neuroinflammation, and gut microbiota to these diseases are highlighted. In this context, the role of specific glial cells (e.g., microglia, astroglia, oligodendrocytes, and synantocytes) on phenotypic manifestation of mood disorders or SUD are emphasized. In addition, use of this knowledge in the potential development of novel therapeutics is touched upon.

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.

Noninvasive photoacoustic computed tomography of mouse brain metabolism in vivo.

We have demonstrated the feasibility of imaging mouse brain metabolism using photoacoustic computed tomography (PACT), a fast, noninvasive and functional imaging modality with optical contrast and acoustic resolution. Brain responses to forepaw stimulations were imaged transdermally and transcranially. 2-NBDG, which diffuses well across the blood-brain-barrier, provided exogenous contrast for photoacoustic imaging of glucose response. Concurrently, hemoglobin provided endogenous contrast for photoacoustic imaging of hemodynamic response. Glucose and hemodynamic responses were quantitatively decoupled by using two-wavelength measurements. We found that glucose uptake and blood perfusion around the somatosensory region of the contralateral hemisphere were both increased by stimulations, indicating elevated neuron activity. While the glucose response area was more homogenous and confined within the somatosensory region, the hemodynamic response area had a clear vascular pattern and spread wider than the somatosensory region. Our results demonstrate that 2-NBDG-enhanced PACT is a promising tool for noninvasive studies of brain metabolism.

Functional significance of cortical NMDA receptors in somatosensory information processing.

N-methyl-d-aspartate receptor (NMDAR)-mediated activity is required for whisker-related neural patterning in the rodent brain. Deletion of the essential NMDAR subunit NR1 gene in excitatory cortical neurons prevents whisker-specific barrel formation and impairs thalamocortical afferent patterning. We used electrophysiological and voltage-sensitive dye imaging methods to assess synaptic and sensory evoked cortical activity and immunohistochemistry to examine immediate early gene expression following whisker stimulation in cortex-specific NR1 knockout (CxNR1KO) mice. In mutant mice, layer IV neurons lacked NMDAR-mediated excitatory postsynaptic currents, and temporal summation of excitatory postsynaptic potentials (EPSPs) was impaired. Barrel neurons showed both phasic and tonic responses to whisker deflection. The averaged tonic response in CxNR1KO mice was significantly less than that in control mice due to impaired EPSP temporal summation. Electrophysiological estimation of the number of thalamic neurons innervating single barrel neurons indicated a significant increase in CxNR1KO mice. Similarly, voltage-sensitive dye optical signals in response to whisker stimulation were widespread. Immediate early gene expression following whisker stimulation also showed a diffuse expression pattern in the CxNR1KO cortex compared with whisker-specific expression patterns in controls. Thus, when NMDAR function is impaired, spatial discrimination of whisker inputs is severely compromised, and sensory stimulation evokes diffuse, topographically misaligned activity in the barrel cortex.

Neurovascular coupling: in vivo optical techniques for functional brain imaging.

Optical imaging techniques reflect different biochemical processes in the brain, which is closely related with neural activity. Scientists and clinicians employ a variety of optical imaging technologies to visualize and study the relationship between neurons, glial cells and blood vessels. In this paper, we present an overview of the current optical approaches used for the in vivo imaging of neurovascular coupling events in small animal models. These techniques include 2-photon microscopy, laser speckle contrast imaging (LSCI), voltage-sensitive dye imaging (VSDi), functional photoacoustic microscopy (fPAM), functional near-infrared spectroscopy imaging (fNIRS) and multimodal imaging techniques. The basic principles of each technique are described in detail, followed by examples of current applications from cutting-edge studies of cerebral neurovascular coupling functions and metabolic. Moreover, we provide a glimpse of the possible ways in which these techniques might be translated to human studies for clinical investigations of pathophysiology and disease. In vivo optical imaging techniques continue to expand and evolve, allowing us to discover fundamental basis of neurovascular coupling roles in cerebral physiology and pathophysiology.

Amyloid Deposition and Dendritic Complexity of Corticocortical Projection Cells in Five Familial Alzheimer's Disease Mouse.

In Alzheimer's disease and related dementias, amyloid beta (Aß) and amyloid plaques can disrupt long-term synaptic plasticity, learning and memory and cognitive function. Plaque accumulation can disrupt corticocortical circuitry leading to abnormalities in sensory, motor, and cognitive processing. In this study, using 5xFAD (five Familial Alzheimer's Disease - FAD - mutations) mice, we evaluated amyloid plaque formation in different cortical areas, and whether differential amyloid accumulation across cortical fields correlates with changes in dendritic complexity of layer 3 corticocortical projection neurons and functional responses in the primary somatosensory cortex following whisker stimulation. We focused on three cortical areas: the primary somatosensory cortex (S1), the primary motor cortex (M1), and the prefrontal cortex (PFC including the anterior cingulate, prelimbic, and infralimbic subdivisions). We found that Aß and amyloid plaque accumulation is not uniform across 5xFAD cortical areas, while there is no expression in littermate controls. We also found that there are differential layer 3 pyramidal cell dendritic complexity changes across the three areas in 5xFAD mice, compared to same age controls, with no apparent relation to differential amyloid accumulation. We used voltage-sensitive dye imaging (VSDi) to visualize neural activity in S1, M1 and PFC following whisker activation. Control mice show normal physiological responses in all three cortical areas, whereas 5xFAD mice only display physiological responses in S1. Taken together our results show that 5xFAD mutation affects the overall dendritic morphology of layer 3 pyramidal cells across sensory-motor and association cortex irrespective of the density and distribution of the Aß amyloid proteins. Corticocortical circuitry between the sensory and motor/association areas is most likely disrupted in 5xFAD mice as cortical responses to whisker stimulation are altered.

In vivo laser speckle contrast imaging of 4-aminopyridine- or pentylenetetrazole-induced seizures.

Clinical and preclinical studies on epileptic seizures are closely linked to the study of neurovascular coupling. Obtaining reliable information about cerebral blood flow (CBF) in the area of epileptic activity through minimally invasive techniques is crucial for research in this field. In our studies, we used laser speckle contrast imaging (LSCI) to gather information about the local blood circulation in the area of epileptic activity. We used two models of epileptic seizures: one based on 4-aminopyridine (4-AP) and another based on pentylenetetrazole (PTZ). We verified the duration of an epileptic seizure using electrocorticography (ECoG). We applied the antiepileptic drug topiramate (TPM) to both models, but its effect was different in each case. However, in both models, TPM had an effect on neurovascular coupling in the area of epileptic activity, as shown by both LSCI and ECoG data. We demonstrated that TPM significantly reduced the amplitude of 4-AP-induced epileptic seizures (4-AP+TPM: 0.61 ± 0.13 mV vs 4-AP: 1.08 ± 0.19 mV; p < 0.05), and it also reduced gamma power in ECoG in PTZ-induced epileptic seizures (PTZ+TPM: 38.5% ± 11.9% of the peak value vs PTZ: 59.2% ± 3.0% of peak value; p < 0.05). We also captured the pattern of CBF changes during focal epileptic seizures induced by 4-AP. Our data confirm that the system of simultaneous cortical LSCI and registration of ECoG makes it possible to evaluate the effectiveness of pharmacological agents in various types of epileptic seizures in in vivo models and provides spatial and temporal information on the process of ictogenesis.

Amount of Melanin Granules in Human Hair Defines the Absorption and Conversion to Heat of Light Energy in the Visible Spectrum.

One of the known important functions of hair is protection from extensive sunlight. This protection is accomplished in large part due to natural hair pigmentation which is known to reflect the number of melanin granules (melanosomes) in the hair shaft, and melanin variants. Melanin takes in excessive light energy and converts it to heat in a process called absorption; heat is then dissipated into the environment as infrared radiation, thereby protecting the underlying skin. We used transmission electron microscopy (TEM) to visualize the melanosome counts in samples of human hair, and used thermal microscopy to measure the temperature changes of the samples when exposed to green and blue light lasers. In our experiments green light conversion to heat was highly correlated to the number of melanosomes, whereas blue light conversion to heat was less correlated, which may be because the reddish melanosomes it contains are less effective in absorbing energy from the blue spectrum of light. Anyway, we have shown the metals accumulation in the melanin can be easily visualized with TEM. We confirmed that the amount of melanin granules in human hair defines the conversion to heat of light energy in the visible spectrum.

Arc protein, a remnant of ancient retrovirus, forms virus-like particles, which are abundantly generated by neurons during epileptic seizures, and affects epileptic susceptibility in rodent models.

A product of the immediate early gene Arc (Activity-regulated cytoskeleton-associated protein or Arc protein) of retroviral ancestry resides in the genome of all tetrapods for millions of years and is expressed endogenously in neurons. It is a well-known protein, very important for synaptic plasticity and memory consolidation. Activity-dependent Arc expression concentrated in glutamatergic synapses affects the long-time synaptic strength of those excitatory synapses. Because it modulates excitatory-inhibitory balance in a neuronal network, the Arc gene itself was found to be related to the pathogenesis of epilepsy. General Arc knockout rodent models develop a susceptibility to epileptic seizures. Because of activity dependence, synaptic Arc protein synthesis also is affected by seizures. Interestingly, it was found that Arc protein in synapses of active neurons self-assemble in capsids of retrovirus-like particles, which can transfer genetic information between neurons, at least across neuronal synaptic boutons. Released Arc particles can be accumulated in astrocytes after seizures. It is still not known how capsid assembling and transmission timescale is affected by seizures. This scientific field is relatively novel and is experiencing swift transformation as it grapples with difficult concepts in light of evolving experimental findings. We summarize the emergent literature on the subject and also discuss the specific rodent models for studying Arc effects in epilepsy. We summarized both to clarify the possible role of Arc-related pseudo-viral particles in epileptic disorders, which may be helpful to researchers interested in this growing area of investigation.

Adjuvant effect of dendritic cells activator Imiquimod in genetic immunization with HIV-1 p55 Gag.

Dendritic cells (DC) are important antigen-presenting cells that have abilities to induce and maintain T-cell immunity, or attenuate it during hyperimmunization. Additional activation of DCs may be useful for vaccination purposes. Imiquimod is known to be a specific agonist of the Toll-like receptors (TLR7), which are located mainly on DCs. To study the effect of DC stimulation on the effectiveness of an HIV-1 p55 gag DNA vaccine in a mice model, we employed 25, 50, and 100 nM of Imiquimod as an adjuvant. Subsequently, Western blot analysis was used to quantify p55 protein production after the immunization. To characterize T-cells immune response, both the frequency of IFN-γ -secreting cells and IFN-γ and IL-4 production were measured, via an ELIspot assay and ELISA, respectively. Low concentrations of Imiquimod were found to effectively stimulate Gag production and the magnitude of the T-cell immune response, whereas higher concentrations reduced vaccination effects. Our results show that the adjuvant effects of Imiquimod depend on concentration. The use of Imiquimod may be helpful to study DC to T cell communication, including possible induction of immunotolerance.

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.

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.

Multilayer subwavelength gratings or sandwiches with periodic structure shape light reflection in the tapetum lucidum of taxonomically diverse vertebrate animals.

Eye shine in the dark has attracted many researchers to the field of eye optics, but the initial studies of subwavelength arrangements in tapetum began only with the development of electronic microscopy at the end of the 20th century. As a result of a number of studies, it was shown that the reflective properties of the tapetum are due to their specialized cellular subwavelength microstructure (photonic crystals). These properties, together with the mutual orientation of the crystals, lead to a significant increase in reflection, which, in turn, enhances the sensitivity of the eye. In addition, research confirmed that optical mechanisms of reflection in the tapetum are very similar even for widely separated species. Due to progress in the field of nano-optics, researchers now have a better understanding of the main principles of this phenomenon. In this review, we summarize electron microscopic and functional studies of tapetal structures in the main vertebrate classes. This allows data on the microstructure of the tapetum to be used to improve our understanding of the visual system.