A paradigm for ethanol consumption in head-fixed mice during prefrontal cortical two-photon calcium imaging.

The prefrontal cortex (PFC) is a hub for cognitive behaviors and is a key target for neuroadaptations in alcohol use disorders. Recent advances in genetically encoded sensors and functional microscopy allow multimodal in vivo PFC activity recordings at subcellular and cellular scales. While these methods could enable a deeper understanding of the relationship between alcohol and PFC function/dysfunction, they typically require animals to be head-fixed. Here, we present a method in mice for binge-like ethanol consumption during head-fixation. Male and female mice were first acclimated to ethanol by providing home cage access to 20% ethanol (v/v) for 4 or 8 days. After home cage drinking, mice consumed ethanol from a lick spout during head-fixation. We used two-photon calcium imaging during the head-fixed drinking paradigm to record from a large population of PFC neurons (>1000) to explore how acute ethanol affects their activity. Drinking exerted temporally heterogeneous effects on PFC activity at single neuron and population levels. Intoxication modulated the tonic activity of some neurons while others showed phasic responses around ethanol receipt. Population level activity did not show tonic or phasic modulation but tracked ethanol consumption over the minute-timescale. Network level interactions assessed through between-neuron pairwise correlations were largely resilient to intoxication at the population level while neurons with increased tonic activity showed higher synchrony by the end of the drinking period. By establishing a method for binge-like drinking in head-fixed mice, we lay the groundwork for leveraging advanced microscopy technologies to study alcohol-induced neuroadaptations in PFC and other brain circuits. This article is part of the Special Issue on "PFC circuit function in psychiatric disease and relevant models".

A paradigm for ethanol consumption in head-fixed mice during prefrontal cortical two-photon calcium imaging.

The prefrontal cortex (PFC) is a hub for higher-level cognitive behaviors and is a key target for neuroadaptations in alcohol use disorders. Preclinical models of ethanol consumption are instrumental for understanding how acute and repeated drinking affects PFC structure and function. Recent advances in genetically encoded sensors of neuronal activity and neuromodulator release combined with functional microscopy (multiphoton and one-photon widefield imaging) allow multimodal in-vivo PFC recordings at subcellular and cellular scales. While these methods could enable a deeper understanding of the relationship between alcohol and PFC function/dysfunction, they require animals to be head-fixed. Here, we present a method in mice for binge-like ethanol consumption during head-fixation. Male and female mice were first acclimated to ethanol by providing home cage access to 20% ethanol (v/v) for 4 or 8 days. After home cage drinking, mice consumed ethanol from a lick spout during head-fixation. We used two-photon calcium imaging during the head-fixed drinking paradigm to record from a large population of PFC neurons (>1000) to explore how acute ethanol affects their activity. Drinking modulated activity rates in a subset of neurons on slow (minutes) and fast (seconds) time scales but the majority of neurons were unaffected. Moreover, ethanol intake did not significantly affect network level interactions in the PFC as assessed through inter-neuronal pairwise correlations. By establishing a method for binge-like drinking in head-fixed mice, we lay the groundwork for leveraging advanced microscopy technologies to study alcohol-induced neuroadaptations in PFC and other brain circuits.

Biodynamic digital holographic speckle microscopy for oocyte and embryo metabolic evaluation.

Assisted reproductive technologies seek to improve the success rate of pregnancies. Morphology scoring is a common approach to evaluate oocyte and embryo viability prior to embryo transfer in utero, but the efficacy of the method is low. We apply biodynamic imaging, based on dynamic light scattering and low-coherence digital holography, to assess the metabolic activity of oocytes and embryos. A biodynamic microscope, developed to image small and translucent biological specimens, is inserted into the bay of a commercial inverted microscope that can switch between conventional microscopy channels and biodynamic microscopy. We find intracellular Doppler spectral features that act as noninvasive proxies for embryo metabolic activity that may relate to embryo viability.

Biodynamic optical assay for embryo viability.

Early stage porcine parthenogenetic embryos were evaluated for metabolic activity using a biodynamic microscope (BDM) that images dynamic light scattering using low-coherence digital holography. The microscope has a 45-deg illumination configuration that reduces specular background for the imaging of small translucent samples. The off-axis illumination is compatible with coherence-gated imaging because of volumetric light scattering in which the coherence plane is tilted at half the illumination angle in a three-dimensional tissue target. The BDM was used to profile the viability of porcine parthenotes with normal and with inhibited mitochondrial adenosine triphosphate (ATP) production using Doppler fluctuation spectroscopy. The ATP concentrations in the parthenotes, which are indicative of developmental potential, were validated by a conventional bioluminescence assay. Biodynamic classifications achieved ∼80 % accuracy correlating sample ATP treatment, providing a quick, label-free surrogate measurement to replace invasive metabolic assays as a candidate for evaluating quality of early embryos in the assisted reproductive technology setting.