The generation and application of human stem-cell-derived functional neural circuits promises novel insights into neurodegenerative diseases. These networks are often studied using stem-cell derived random neural networks in vitro, with electrical stimulation and recording using multielectrode arrays. However, the impulse response function of networks is best obtained with spatiotemporally well-defined stimuli, which electrical stimulation does not provide. Optogenetics allows for the functional control of genetically altered cells with light stimuli at high spatiotemporal resolution. Current optogenetic investigations of neural networks are often conducted using full field illumination, potentially masking important functional information. This can be avoided using holographically shaped illumination. In this article, we present a digital holographic illumination setup with a spatial resolution of about 8 µm, which suffices for the stimulation of single neurons, and offers a temporal resolution of less than 0.6 ms. With this setup, we present preliminary single-cell stimulation recording of stem-cell derived induced human neurons in a random neural network. This will offer the opportunity for further studies on connectivity in such networks.

Laser ultrasonics is a powerful technique for contactless investigation of important material parameters such as Young’s modulus or thin layer thickness. However, the often employed Gaussian beams result in diverging sound fields of quickly decreasing intensity. Conventionally, changing the laser beam profile requires the slow movement or exchange of optical elements. We present a laser ultrasonics setup for the creation of arbitrary intensity distributions by holographic projection using a MEMS spatial light modulator. High-intensity ultrasound foci with a focus width of 1.6 mm are scanned axially in a sample into depths of up to 7.4 mm by projecting ring-shaped intensity distributions of varying diameter without any mechanical movements. This technique is promising for highly spatially resolved flaw detection or a fast scanning investigation of biological tissue.

Optogenetics is a versatile toolset for the functional investigation of excitable cells such as neurons and cardiomyocytes in vivo and in vitro. While monochromatic illumination of these cells for either stimulation or inhibition already enables a wide range of studies, the combination of activation and silencing in one setup facilitates new experimental interrogation protocols. In this work, we present a setup for the simultaneous holographic stimulation and inhibition of multiple cells in vitro. The system is based on two fast ferroelectric liquid crystal spatial light modulators with frame rates of up to 1.7 kHz. Thereby, we are able to illuminate up to about 50 single spots with better than cellular resolution and without crosstalk, perfectly suited for refined network analysis schemes. System-inherent aberrations are corrected by applying an iterative optimization scheme based on Zernike polynomials. These are superposed on the same spatial light modulators that display the pattern-generating holograms, hence no further adaptive optical elements are needed for aberration correction. A near-diffraction-limited spatial resolution is achieved over the whole field of view, enabling subcellular optogenetic experiments by just choosing an appropriate microscope objective. The setup can pave the way for a multitude of optogenetic experiments, in particular with cardiomyocytes and neural networks.

HiPSCNN Functional connectivity was investigated using optogenetics. Time-distance-correlations can be obtained using holographic single-neuron stimulation and provide deeper insights into of structure-function relations than full-field stimulation.

A two-wavelength optogenetic stimulation platform with high spatiotemporal resolution and inherent aberration correction is presented. By stimulating single neurons, we investigated the temporal evolution of connectivity in human induced pluripotent stem cell-derived neuronal networks in-vitro.