Transparent neural interfaces: challenges and solutions of microengineered multimodal implants designed to measure intact neuronal populations using high-resolution electrophysiology and microscopy simultaneously.

The aim of this review is to present a comprehensive overview of the feasibility of using transparent neural interfaces in multimodal in vivo experiments on the central nervous system. Multimodal electrophysiological and neuroimaging approaches hold great potential for revealing the anatomical and functional connectivity of neuronal ensembles in the intact brain. Multimodal approaches are less time-consuming and require fewer experimental animals as researchers obtain denser, complex data during the combined experiments. Creating devices that provide high-resolution, artifact-free neural recordings while facilitating the interrogation or stimulation of underlying anatomical features is currently one of the greatest challenges in the field of neuroengineering. There are numerous articles highlighting the trade-offs between the design and development of transparent neural interfaces; however, a comprehensive overview of the efforts in material science and technology has not been reported. Our present work fills this gap in knowledge by introducing the latest micro- and nanoengineered solutions for fabricating substrate and conductive components. Here, the limitations and improvements in electrical, optical, and mechanical properties, the stability and longevity of the integrated features, and biocompatibility during in vivo use are discussed.

Transparent, low-autofluorescence microECoG device for simultaneous Ca2+ imaging and cortical electrophysiology in vivo.

OBJECTIVE: Multimodal neuroimaging approaches are beneficial to discover brain functionalities at high spatial and temporal resolution. In our work, a novel material composition of a microECoG device relying on Parylene HT and indium-tin-oxide (ITO) is presented, which facilitates two-photon imaging of Ca2+ signals and concurrent recording of cortical EEG. APPROACH: Long-term stability of the interfaces of the transparent microdevice is confirmed in vitro by electrochemical and mechanical tests. The outstanding optical properties, like high transmittance and low auto-fluorescent are proven by fluorimetric measurements. Spatial resolution of fluorescent two-photon imaging through the microECoG device is presented in transgenic hippocampal slices, while concurrent recording of Ca2+ signals and cortical EEG is demonstrated in vivo. Photoartefacts and photodegradation of the materials are also investigated in detail to provide safety guidelines for further use in two-photon in vivo imaging schemes. MAIN RESULTS: Two-photon imaging of Ca signals can be safely performed through the proposed transparent ECoG device, without significant distortion in the dimensions of detected neuronal structures or in the temporal signaling. In chronic use, we demonstrated that fluorescent Ca signals of individual neurons can be clearly recorded even after 51 d. SIGNIFICANCE: Our results give a firm indication that highly transparent microECoG electrode arrays made of Parylene HT/ITO/Parylene HT multilayer are excellent candidates for synergetic recording of optical signals and EEG from intact brains with high resolution and are free of electrical and optical artefacts.

Neuronal and synaptic structure of the specific thalamic nuclei.

1. Neuronal organization of the specific thalamic nuclei is rather uniform. The ratio of thalamocortical relay neuron and Golgi II type interneuron is 2:1, as well as 3:1. 2. Interneurons show GABA-immunoreactivity. The presynaptic dendrites (F2-profiles) modify the intrathalamic transmission of the specific afferent impulses. 3. The two principal types of synaptic arrangements were observed in the specific thalamic nuclei: the synaptic glomeruli and the general neuropil. 4. Three different types of axon-terminals could be distinguished in the neuropil of all specific thalamic nuclei: the large RL-boutons are terminals of the specific afferents, the small RS-boutons are mainly the terminals of cortical afferents and the F1-boutons are probably axon-terminals of the Golgi II type interneurons, as well as axon-endings originating from the reticular nucleus of thalamus. 5. Synaptic contacts of the specific afferent fibers have basic importance in the relay nuclei. 6. In associative and anterior thalamic nuclei the RS- and F1-boutons have a significant modification effect on the interneurons (inhibition-disinhibition).

[Limits on the use of propylene glycol in topical preparations]. / Limites d'utilisation du propylène glycol dans les produits à usage topique.

The authors define some limits for useful concentrations of propylene glycol (P.G.) by determination of inhibition produced in cell respiration from (crushed) mouse liver and in human cutaneous fibroblastic proliferation. They conclude that P.G. is able to inhibit both these physicological mechanisms from concentration as weak as 5.5% and 2% respectively. They give a method for analytical determination of P.G. in cosmetic and pharmaceutical products using gas liquid chromatography. This method is useful for a wide range of concentrations.

Morphological alterations in subcortical vibrissal relays following vibrissal follicle destruction at birth in the mouse.

Morphological modifications of two subcortical vibrissal relays were analyzed, following destruction of vibrissal follicles in newborn mice. The volume of the nucleus interpolaris (NI) of the trigeminal nuclear complex in the brainstem decreased by 33%, while the number of its neuronal perikarya decreased only moderately. Vibrissal deafferentation caused no shrinkage of the ventrobasal complex (VB). In the damaged medial vibrissal part of VB (VBm), however, neuronal density was higher than normal, indicating the prevention or retardation of physiologically programmed cell death in the afferentation deprived thalamic somatosensory relay station. It is suggested that the difference in neuron density produced by deafferentation is related to the states of maturation at birth of the two subcortical vibrissal relays. Following vibrissal deafferentation the basic organization of the synaptic neuropil appeared to be similar to the control. Quantitative electron microscopic (EM) analysis revealed, however, an increased number of axon terminals with ovoid synaptic vesicles in both deafferented relay stations. The increased density of gamma-aminobutyric acid (GABA)-immunostained boutons observed in the VBm following vibrissal deprivation suggested a compensatory increase most probably of the inhibitory axon endings. Quantitative EM analysis also provided evidence that many or most of the specific afferent terminals in the damaged VBm were not identical with but were substitutes for the original "vibrissal" specific afferents. Forty percent of all "specific" afferents were shown to be modified corticothalamic terminals. The modification and the resemblence of some cortical endings to specific afferents demonstrated the morphogenetic plasticity of synaptogenesis in these terminals during development as well as the importance and inductive potential of the postsynaptic target in the differentiation of presynaptic axon terminals.

Numerical estimation of gamma-aminobutyric acid (GABA)-containing neurons in three thalamic nuclei of the cat: direct GABA immunocytochemistry.

The percentage of neurons that are immunoreactive for the inhibitory neurotransmitter, gamma-aminobutyric acid (GABA) was determined within: (1) the lateral geniculate nucleus (LGN), (2) the ventrobasal complex (VB) and (3) the antero-ventral nucleus (AV) of the thalamus in the cat. An antiserum to GABA was used to stain GABA-containing perikarya in 1.0 micrometer thick Araldite-embedded sections. Immunostained somata in all three nuclei were invariably smaller than the immuno-negative nerve cells. 27% of all neurons in the LGN, 33% in the VB and 25% in the AV were immunoreactive for GABA.

[Histological changes in the somatosensory thalamus after unilateral coagulation of vibrissa follicles of the muzzle of the newborn mouse]. / Modifications histologiques dans le thalamus somatosensoriel après coagulation unilatérale des follicules des vibrisses du museau chez la Souris nouveaunée.

Light microscopic study of the thalamic ventro-basal complex (VB), after unilateral coagulation of vibrissae follicles in newborn mouse, revealed an excess of neuronal perikarya on the controlateral "deafferented" side as compared to the normal side. The higher density of nerve cells was confined to the vibrissal relay in the medial part of VB nucleus (VBm), whereas the cell number in the non vibrissal-lateral part of this nucleus (VB1) remained on the control level. Electron microscopic investigation of the thalamic vibrissal relay has shown signs of a modified synaptogenesis on the "deafferented" side: (a) the number of specific afferents has diminished and in contrast to the normal side, most of the specific sensory terminals contain small spheroid synaptic vesicles and (b) the number of axon terminals with ovoid pleomorphic vesicles has been doubled.

Quantitative aspects in the analysis of the synaptic architecture of thalamic sensory relay nuclei.

Special orientations of dendritic trees of projective (relay) neurons were observed in the thalamic nuclei; and it was found to be different in the various nuclei. It can be explained by the size, course, site of the fibre bundles which enter and/or transgress the nuclei. The orientation of dendritic tree and the size of the branching area of dendrites (specific active dendritic space) were analysed by computer. The quantitative data of neuronal elements in some thalamic nuclei gave the opportunity to consider the possible degree of divergence and convergence of sensory fibres regarding their connections with the projective neurons.

Quantitative histological study on the thalamic ventro-basal complex of the cat. Numerical aspects of the transfer from medial lemniscal fibers to cortical relay.

A quantitative analysis was carried out on the thalamic ventrobasal (VB) complex of the cat. The following numerical and metrical parameters of the neuronal elements (cells and fibers) were determined: 1. Volume of nucleus: 27.38 mm3 2. Total number of neurons: 243,000 3. Total number of fibers of medial lemniscus: 26,000 4. Volume of arborization space of one lemniscal fiber: 2.26 X 10(6) micrometers 3. Numerical data of relay neurons and lemniscal fibers and their relations as basic factors in the estimation of the degree of divergence and convergence of lemniscal input were calculated and compared. It was found that the probable degree of convergence is four-fold (1-4) and of divergence 27-fold (1-27) with regard to the relationship of fibers and cells in the VB. The quantitative data obtained in the VB and our considerations on convergence and divergence were compared with analogous values obtained for the lateral geniculate body LGB. The differences between the two sensory relay nuclei reflect differences in their modes of impulse transmission.

Residual neurons in the lateral geniculate nucleus of adult cats following chronic disconnection from the cortex.

Residual neurons following chronic surgical disconnection from the cortex were studied by light and electron microscopy in the dorsal Lateral Geniculate Nucleus (LGN) of adult cats. One year after operation the volume of the decorticated LGN had shrunken to approximately half that of the control LGN. The number of nerve cells decreased at the same time, to 13-15% of the control, with relatively higher cell loss in A and A1 laminae than in lamina C. The Golgi, Golgi-EM and EM analysis of the residual neurons revealed that they contain two distinct nerve cell types. (1) 55% of all surviving neurons were identified as geniculo-cortical relay cells, while (2) 45% of the persisting nerve cells were interneurons. These data suggest that in the normal LGN of adult cat 7% (or more) of all nerve cells are local interneurons. Finally, those factors which might contribute to the unexpected survival of many relay neurons to the axotomy-caused retrograde degeneration, are considered and discussed.

A combined horseradish peroxidase and Golgi study on the afferent connections of the ventrobasal complex of the thalamus in the cat.

Afferent connections to the ventrobasal complex (VB) of the thalamus were studied by means of retrograde transport of horseradish peroxidase (HRP) and by the Golgi-method. After HRP-injection into the VB, peroxidase-positive cells were observed contralaterally in the dorsal column nuclei (DCN), in the trigeminal nuclei and in the lateral cervical nucleus (LCN), and ipsilaterally in the somatosensoty I (SI) and II (SII) cortical areas. Labeled cells of different shape and size were compared with neurons impregnated by the Golgi-technique. On tn size and shape even within one region and that they correspond to the relay or efferent neurons observed in the Golgi-material.

Types of thalamo-cortical relay neurons in the anteroventral nucleus of the cat. A combined horseradish peroxidase--Golgi study.

Neurons displaying a thalamo-cortical projection were marked by means of the retrograde transport of horseradish peroxidase (HRP), and the labeled elements were compared with neurons impregnated by the Golgi technique. Injections of HRP into the posterior area of the limbic cortex resulted in its uptake by various anterior thalamic nuclei, especially the anteroventral nucleus. HRP-positive cells are characterized by their position, dendritic orientation, and the shape and size of their somata. On the basis of the combined HRP-and Golgi-analysis three different types of thalamo-cortical relay neurons can be distinguished.

Quantitative histological studies on the lateral geniculate nucleus in the cat. III. Distribution of different types of neurons in the several layers of LGN.

An attempt is made on the definition of proportional distribution of the different types of neurons in the layers of LGN by means of quantitative method. On the basis of nuclear volume, nuclear and cell diameters the different types of neurons were separated according to their size and shape. The ratio of relay and interneurons proved to be 2:1 in all layers of LGN of cat. The proportional distribution of further subgroups of neurons was also defined. The bilateral connections between the relay and interneurons based on initial axon collateral of TCR neurons and on axon terminals and dendritic appendages of INs, are also calculated. The interneurons--in the LGN--have widespread contacts with both types of relay neurons, similarly the relay neurons contact the interneurons. The numerical data of morphological observations are accord with the results of physiologists.

Limbic projections to the cat thalamus. A horseradish peroxidase study.

Retrograde axonal transport of horseradish peroxidase was used to identify the afferent projection systems of the anterior thalamic nuclei in the cat. The results indicate that fibres projecting to the three anterior nuclei of the thalamus arise from the hippocampal region, septum and cingulate cortex unilaterally and from the hypothalamus (mamillary nuclei) both bilaterally and unilaterally. The cells of the medial mamillary nuclei send their axons to the ipsilateral AV and AM nuclei whereas the lateral mamillary nucleus projects bilaterally to the AD nuclei.

Granule cells, mossy fibres and pyramidal neurons: an electron microscopic study of the cat's hippocampal formation, I.

The ultrastructure of mossy fibre terminals, granule cells, and pyramidal neurons was studied in cat material. The average of the mossy terminals and the density of the synaptic vesicles were analysed. The large mossy terminals establish mainly axo-spiny synapses on the apical dendrites of pyramidal neurons in the CA3 area, but some axo-dendritic synapses were also observed. Mossy fibre endings were found in the subgranular layer, and also on the dendrites of interneurons.

Quantitative histological studies on the lateral geniculate nucleus in the cat. IV. Numerical aspects of the transfer from restinal fibers to cortical relay.

The numerical data, gained both from the quantitative histology and Golgi stained material was completed with those of optic fibers. The number of X and Y fibers was estimated on the basis of our results, on the data of literature and from the ratio of the large and medium size relay neurons. The crossed and uncrossed fibers were counted and compared with the number of relay neurons in layer A and A1. Knowing the different numerical and metrical counts of neuronal elements in LGN of cat some calculations were carried out on the overlaping of optic fiber arborizations, on the intersections of dendritic ramification points of relay neurons and on the spatial arrangement of both the optic terminals and the relay neurons. The comparison of the neumerical data on relay neurons and optic fibers and their relations were the basic factors of considerations on divergence and convergence of retinal input.

Quantitative histological studies on the lateral geniculate nucleus in the cat. I. Measurements on Golgi material.

Numerical data of neuron and fiber elements were observed and measured on Golgi preparations. The aim of the quantitative Golgi analysis was to obtain data on the spatial arrangement of arely neurons, their relations to the interneurons, on the spatial architecture of the arborizations of optic fibers and their relations to the main neuron types. The quantitative data on arely neurons include numbers of dendrites, of their chief points of ramification as the main targets of the optic afferents, geometric data on the "specific active sphere" of the dendritic tree; on the arborization of initial collaterals. The data on interneurons refer both to numeric and geometric parameters of their axonal arborizations, and to those of various characteristics of their dendritic processes. The data on optic fibers include the size of their arborization space, the numbers of bouton clusters and preterminal bouquets per arborization.

Quantitative histological studies on the lateral geniculate nucleus in the cat. II. Cell numbers and densities in the several layers.

In the cat lateral geniculate nucleus all cells of all types were counted: their absolute numbers, their densities and their relative distribution. The nuclear volume and the volume of each layer was measured. The data of volume was applied on living material. The average densities of cells were calculated from the volume of LGN. The density of the cells was combined with the "specific active dendritic sphere" of relay neurons and it could be estimated that there may be a large number of spatially overlaping of ramification points around the cell body of relay neurons.