The McGill-Mouse-Miniscope platform: A standardized approach for high-throughput imaging of neuronal dynamics during behavior

Understanding the rules that govern neuronal dynamics throughout the brain to subserve behavior and cognition remains one of the biggest challenges in neuroscience research. Recent technical advances enable the recording of increasingly larger neuronal populations to produce increasingly more sophisticated datasets. Despite bold and important open-science and data-sharing policies, these datasets tend to include unique data acquisition methods, behaviors, and file structures. Discrepancies between experimental protocols present key challenges in comparing data between laboratories and across different brain regions and species. Here, we discuss our recent efforts to create a standardized and high-throughput research platform to address these issues. The McGill-Mouse-Miniscope (M3) platform is an initiative to combine miniscope calcium imaging with standardized touchscreen-based animal behavioral testing. The goal is to curate an open-source and standardized framework for acquiring, analyzing, and accessing high-quality data of the neuronal dynamics that underly cognition throughout the brain in mice, marmosets, and models of disease. We end with a discussion of future developments and a call for users to adopt this standardized approach.     

FKBP12.6 overexpression decreases Ca2+ spark amplitude but enhances [Ca2+]i transient in rat cardiac myocytes

Ryanodine receptors/Ca2+-release channels (RyR2) from the sarcoplasmic reticulum (SR) provide the Ca2+ required for contraction at each cardiac twitch. RyR2 are regulated by a variety of proteins, including the immunophilin FK506 binding protein (FKBP12.6). FKBP12.6 seems to be important for coupled gating of RyR2 and its deficit and alteration may be involved in heart failure. The role of FKBP12.6 on Ca2+ release has not been analyzed directly, but rather it was inferred from the effects of immunophilins, such us FK506 and rapamycin, which, among other effects, dissociates FKBP12.6 from the RyR2. Here, we investigated directly the effects of FKBP12.6 on local (Ca2+ sparks) and global [intracellular Ca2+ concentration ([Ca2+]i) transients] Ca2+ release in single rat cardiac myocytes. The FKBP12.6 gene was transfected in single myocytes using the adenovirus technique with a reporter gene strategy based on green fluorescent protein (GFP) to check out the success of transfections. Control myocytes were transfected with only GFP (Ad-GFP). Rhod-2 was used as the Ca2+ indicator, and cells were viewed with a confocal microscope. We found that overexpression of FKBP12.6 decreases the occurrence, amplitude, duration, and width of spontaneous Ca2+ sparks. FK506 had diametrically opposed effects. However, overexpression of FKBP12.6 increased the [Ca2+]i transient amplitude and accelerated its decay in field-stimulated cells. The associated cell shortening was increased. SR Ca2+ load, estimated by rapid caffeine application, was increased. In conclusion, FKBP12.6 overexpression decreases spontaneous Ca2+ sparks but increases [Ca2+]i transients, in relation with enhanced SR Ca2+ load, therefore improving excitation-contraction coupling.     

Polynomial algebra reveals diverging roles of the unfolded protein response in endothelial cells during ischemia–reperfusion injury

The unfolded protein response (UPR) – the endoplasmic reticulum stress response – is found in various pathologies including ischemia–reperfusion injury (IRI). However, its role during IRI is still unclear. Here, by combining two different bioinformatical methods – a method based on ordinary differential equations (Time Series Network Inference) and an algebraic method (probabilistic polynomial dynamical systems) – we identified the IRE1α–XBP1 and the ATF6 pathways as the main UPR effectors involved in cell’s adaptation to IRI. We validated these findings experimentally by assessing the impact of their knock-out and knock-down on cell survival during IRI.     

Unique properties of polyphenol stilbenes in the brain: more than direct antioxidant actions; gene/protein regulatory activity

The 'French Paradox' has been typically associated with moderate consumption of wine, especially red wine. A polyphenol 3,4',5-trihydroxy-trans-stilbene (a member of the non-flavonoids family), better known as resveratrol, has been purported to have many health benefits. A number of these valuable properties have been attributed to its intrinsic antioxidant capabilities, although the potential level of resveratrol in the circulation is likely not enough to neutralize free radical scavenging. The brain and the heart are uniquely vulnerable to hypoxic conditions and oxidative stress injuries. Recently, evidence suggests that resveratrol could act as a signaling molecule within tissues and cells to modulate the expression of genes and proteins. Stimulation of such proteins and enzymes could explain some the intracellular antioxidative properties. The modulation of genes could suffice as an explanation of some of resveratrol's cytoprotective actions, as well as its influence on blood flow, cell death, and inflammatory cascades. Resveratrol stimulation of the expression of heme oxygenase is one example. Increased heme oxygenase activity has led to significant protection against models of in vitro and in vivo oxidative stress injury. Resveratrol could provide cellular resistance against insults; although more work is necessary before it is prescribed as a potential prophylactic in models of either acute or chronic conditions, such as stroke, amyotrophic lateral sclerosis, Parkinson, Alzheimer, and a variety of age-related vascular disorders.     

Environmental factors influencing the regional distribution and local density of a small benthic fish: the stoneloach (<Emphasis Type="Italic">Barbatula barbatula</Emphasis>)

We investigated the relationships between different environmental variables and the spatial distribution patterns of the stoneloach (Barbatula barbatula) at the stream system, the stream site, and the mesohabitat (riffle/pool) scales in south-western France. Stoneloach occurred at 240 sites (out of 554 sampling sites), chiefly close to the source, in areas at low elevation and with weak slopes. Population density at a site was primarily influenced by physical conditions. Stream width was positively related to the probability of presence of stoneloach within the stream system, but negatively related to local density. These results indicate that stoneloaches can occur in a wide range of streams, but they are less abundant in wide rivers, probably because of lower habitat heterogeneity. Slope was negatively correlated to both fish presence at the regional scale and local density, suggesting that stoneloach’s swimming performance were weak under greater erosive forces. These results suggested that the distribution of populations and the density of stoneloach were governed by the suitability of physical habitat. Multi-scale studies of factors influencing a species’distribution allow to integrate patterns observed at different scales, and enhance our understanding of interactions between animals and their environment. The use of few pertinent variables in successful final models could reduce the effort and cost of data collection for water management applications.     

Anti-PrP antibodies block PrPSc replication in prion-infected cell cultures by accelerating PrPC degradation

The use of anti-PrP antibodies represents one of the most promising strategies for the treatment of prion diseases. In the present study, we screened various anti-PrP antibodies with the aim of identifying those that would block PrP(Sc) replication in prion-infected cell culture. Two antibodies, SAF34 recognizing the flexible octarepeats region on HuPrP protein, and SAF61 directed against PrP amino acid residues (144-152), not only inhibited PrP(Sc) formation in prion-infected neuroblastoma cells but also decreased the PrP(C) levels in non-infected N2a cells. In addition, treatment with both SAF34 and SAF61 antibodies decreased PrP(C) and PrP(Sc) levels in the cells synergistically. In the presence of both antibodies, our results showed that the mode of action which leads to the disappearance of PrP(Sc) in cells is directly coupled to PrP(C) degradation by reducing the half-life of the PrP(C) protein.     

X-linked mental retardation and autism are associated with a mutation in the NLGN4 gene, a member of the neuroligin family

A large French family including members affected by nonspecific X-linked mental retardation, with or without autism or pervasive developmental disorder in affected male patients, has been found to have a 2-base-pair deletion in the Neuroligin 4 gene (NLGN4) located at Xp22.33. This mutation leads to a premature stop codon in the middle of the sequence of the normal protein and is thought to suppress the transmembrane domain and sequences important for the dimerization of neuroligins that are required for proper cell-cell interaction through binding to beta-neurexins. As the neuroligins are mostly enriched at excitatory synapses, these results suggest that a defect in synaptogenesis may lead to deficits in cognitive development and communication processes. The fact that the deletion was present in both autistic and nonautistic mentally retarded males suggests that the NLGN4 gene is not only involved in autism, as previously described, but also in mental retardation, indicating that some types of autistic disorder and mental retardation may have common genetic origins.