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Abstract In mammals, the suprachiasmatic nucleus of the hypothalamus is the master circadian pacemaker that synchronizes the clocks in the central nervous system and periphery, thus orchestrating rhythms throughout the body. However, little is known about how so many cellular clocks within and across brain circuits can be effectively synchronized. In this work, we investigated the implication of two possible pathways: (i) astrocytes-mediated synchronization and (ii) neuronal paracrine factors-mediated synchronization. By taking advantage of a lab-on-a-chip microfluidic device developed in our laboratory, here we report that both pathways are involved. We found the paracrine factors-mediated synchronization of molecular clocks is diffusion-limited and, in our device, effective only in case of a short distance between neuronal populations. Interestingly, interconnecting astrocytes define an active signaling channel that can synchronize molecular clocks of neuronal populations also at longer distances. At mechanism level, we found that astrocytes-mediated synchronization involves both GABA and glutamate, while neuronal paracrine factors-mediated synchronization occurs through GABA signaling. These findings identify a previously unknown role of astrocytes as active cells that might distribute long-range signals to synchronize the brain clocks, thus further strengthening the importance of reciprocal interactions between glial and neuronal cells in the context of circadian circuitry.

Flexible polymer neural probes are an attractive emerging approach for invasive brain recordings, given that they can minimize the risks of brain damage or glial scaring. However, densely packed electrode sites, which can facilitate neuronal data analysis, are not widely available in flexible probes. Here, we present a new flexible polyimide neural probe, based on standard and low-cost lithography processes, which has 32 closely spaced 10 μm diameter gold electrode sites at two different depths from the probe surface arranged in a matrix, with inter-site distances of only 5 μm. The double-layer design and fabrication approach implemented also provides additional stiffening just sufficient to prevent probe buckling during brain insertion. This approach avoids typical laborious augmentation strategies used to increase flexible probes’ mechanical rigidity while allowing a small brain insertion footprint. Chemical composition analysis and metrology of structural, mechanical, and electrical properties demonstrated the viability of this fabrication approach. Finally, in vivo functional assessment tests in the mouse cortex were performed as well as histological assessment of the insertion footprint, validating the biological applicability of this flexible neural probe for acquiring high quality neuronal recordings with high signal to noise ratio (SNR) and reduced acute trauma.

Minimally invasive medical devices can greatly benefit from Narrow Band Imaging (NBI) diagnostic capabilities, as different wavelengths allow penetration of distinct layers of the gastrointestinal tract mucosa, improving diagnostic accuracy and targeting different pathologies. An important performance parameter is the light intensity at a given power consumption of the medical device. A method to increase the illumination intensity in the NBI diagnostic technique was developed and applied to minimally invasive medical devices (e.g., endoscopic capsules), without increasing the size and power consumption of such instruments. Endoscopic capsules are generally equipped with light-emitting diodes (LEDs) operating in the RGB (red, green, and blue) visible light spectrum. A polydimethylsiloxane (PDMS) µ-lens was designed for a maximum light intensity at the target area of interest when placed on top of the LEDs. The PDMS µ-lens was fabricated using a low-cost hanging droplet method. Experiments reveal an increased illumination intensity by a factor of 1.21 for both the blue and green LEDs and 1.18 for the red LED. These promising results can increase the resolution of NBI in endoscopic capsules, which can contribute to early gastric lesions diagnosis.

Neural probes can be equipped with light for optogenetics applications. Different approaches are used for delivering light to the tissue: an optical fiber coupled to the probe, a µLED or a waveguide integrated on the probe. Small probe dimensions, adequate optical power for photostimulation and good tissue penetration for in-vivo experiments are critical requirements. Thus, integrating a waveguide is a promising solution. This work shows the design and simulation of a SU-8 based waveguide for integration in a neural probe. The waveguide contains 3 apertures, spaced by 0.5 mm, which will allow the photostimulation of different brain regions simultaneously.

In optogenetic studies, the brain is exposed to high-power light sources and inadequate power density or exposure time can cause cell damage from overheating (typically temperature increasing of 2 ∘ C). In order to overcome overheating issues in optogenetics, this paper presents a neural tool capable of assessing tissue temperature over time, combined with the capability of electrical recording and optical stimulation. A silicon-based 8 mm long probe was manufactured to reach deep neural structures. The final proof-of-concept device comprises a double-sided function: on one side, an optrode with LED-based stimulation and platinum (Pt) recording points; and, on the opposite side, a Pt-based thin-film thermoresistance (RTD) for temperature assessing in the photostimulation site surroundings. Pt thin-films for tissue interface were chosen due to its biocompatibility and thermal linearity. A single-shaft probe is demonstrated for integration in a 3D probe array. A 3D probe array will reduce the distance between the thermal sensor and the heating source. Results show good recording and optical features, with average impedance magnitude of 371 k Ω , at 1 kHz, and optical power of 1.2 mW·mm − 2 (at 470 nm), respectively. The manufactured RTD showed resolution of 0.2 ∘ C at 37 ∘ C (normal body temperature). Overall, the results show a device capable of meeting the requirements of a neural interface for recording/stimulating of neural activity and monitoring temperature profile of the photostimulation site surroundings, which suggests a promising tool for neuroscience research filed.

Advancements in stem cell technology together with an improved understanding of in vitro organogenesis have enabled new routes that exploit cell-autonomous self-organization responses of adult stem cells (ASCs) and homogenous pluripotent stem cells (PSCs) to grow complex, three-dimensional (3D), mini-organ like structures on demand, the so-called organoids. Conventional optical and electrical neurophysiological techniques to acquire functional data from brain organoids, however, are not adequate for chronic recordings of neural activity from these model systems, and are not ideal approaches for throughput screenings applied to drug discovery. To overcome these issues, new emerging approaches aim at fusing sensing mechanisms and/or actuating artificial devices within organoids. Here we introduce and develop the concept of the Lab-in-Organoid (LIO) technology for in-tissue sensing and actuation within 3D cell aggregates. This challenging technology grounds on the self-aggregation of brain cells and on integrated bioelectronic micro-scale devices to provide an advanced tool for generating 3D biological brain models with in-tissue artificial functionalities adapted for routine, label-free functional measurements and for assay’s development. We complete previously reported results on the implementation of the integrated self-standing wireless silicon micro-devices with experiments aiming at investigating the impact on neuronal spheroids of sinusoidal electro-magnetic fields as those required for wireless power and data transmission. Finally, we discuss the technology headway and future perspectives.