Nondestructive evaluation of progressive neuronal changes in organotypic rat hippocampal slice cultures using ultrahigh-resolution optical coherence microscopy

Abstract

Three-dimensional tissue cultures have been used as effective models for studying different diseases including epilepsy. High-throughput, non-destructive techniques are essential for rapid assessment of disease-related processes such as progressive cell death. In this study, an ultrahigh-resolution optical coherence microscopy (UHR-OCM) system with ~1.5-μm axial resolution and ~2.3-μm transverse resolution was developed to evaluate seizure-induced neuronal injury in organotypic rat hippocampal cultures. The capability of UHR-OCM to visualize cells in neural tissue was confirmed by comparison of UHR-OCM images with confocal immunostained images of the same cultures. In order to evaluate the progression of neuronal injury, UHR-OCM images were obtained from cultures on 7, 14, 21 and 28 days-in-vitro (DIVs). In comparison to DIV 7, statistically significant reductions in 3D cell count and culture thickness from UHR-OCM images were observed on subsequent time points. In cultures treated with kynurenic acid (KYNA), significantly less reduction in cell count and culture thickness was observed compared to control specimens. These results demonstrate the capability of UHR-OCM to perform rapid, label-free, and non-destructive evaluation of neuronal death in organotypic hippocampal cultures. UHR-OCM, in combination with three-dimensional tissue cultures, can potentially prove to be a promising tool for high throughput screening of drugs targeting various disorders.

Optical Coherence Tomography for Brain Imaging and Developmental Biology

“…OCM revealed the same individual neurons observed in confocal microscopy (Fig. 1C, D) [78], two-photon microscopy [34, 77] and histological slides [79, 80]. Neuron count obtained with OCM from 3D organotypic brain cultures also showed linear correlation with confocal microscopy (Fig.…”

“…Neuron count obtained with OCM from 3D organotypic brain cultures also showed linear correlation with confocal microscopy (Fig. 1E) [78]. …”

“…OCM is able to resolve individual neurons based on intrinsic optical contrast in rodent brains [34, 77, 78] and in human brain slices [79, 80]. Neurons are shown as hyposcattering regions in OCM images (Fig.…”

Wide-field optical coherence microscopy of the mouse brain slice

“…OCM has been widely researched for visualizing single myelin fiber [2], cerebral cortex [3], tumorous tissue [4], laminar structure of isocortex [5], and neuronal cell bodies in the brain [6,7]. Whereas most OCM systems provide the regional geometric anatomy with narrow field of view, so it has restriction to derive important information such as the microscopic feature of myelin and their overall connectivity [8].…”

Optical Coherence Tomography Detects Necrotic Regions and Volumetrically Quantifies Multicellular Tumor Spheroids

“…We have recently developed high resolution OCT imaging platforms for various in vitro bioapplications, including examining angiogenic sprouting in microengineered devices (43) and evaluating progressive neuronal changes in in vitro rat hippocampal slice cultures over time (44). In this study, we employed a label-free OCT-based imaging platform to visualize 3D structures of multicellular tumor spheroids (MCTSs) in 96-well plates and obtained high resolution structural and physiological information from the spheroids over a period of 21 days.…”