Psychedelics and neural plasticity.

When it comes to studying neural plasticity and psychedelics, the numerous and diverse neuroscientific fields converging on the topic provide unique insight into a complex picture. This editorial will describe the major ways in which the known effects of psychedelics on plasticity are being studied. We lay out strengths of different techniques and the major gaps and room for future research, particularly in the translation of pre-clinical studies to human research.

Phosphorylation of PSD-95 at serine 73 in dCA1 is required for extinction of contextual fear.

The updating of contextual memories is essential for survival in a changing environment. Accumulating data indicate that the dorsal CA1 area (dCA1) contributes to this process. However, the cellular and molecular mechanisms of contextual fear memory updating remain poorly understood. Postsynaptic density protein 95 (PSD-95) regulates the structure and function of glutamatergic synapses. Here, using dCA1-targeted genetic manipulations in vivo, combined with ex vivo 3D electron microscopy and electrophysiology, we identify a novel, synaptic mechanism that is induced during attenuation of contextual fear memories and involves phosphorylation of PSD-95 at Serine 73 in dCA1. Our data provide the proof that PSD-95-dependent synaptic plasticity in dCA1 is required for updating of contextual fear memory.

Stressors Length and the Habituation Effect-An EEG Study.

The research described in this paper aimed to determine whether people respond differently to short and long stimuli and whether stress stimuli repeated over time evoke a habituation effect. To meet this goal, we performed a cognitive experiment with eight subjects. During this experiment, the subjects were presented with two trays of stress-inducing stimuli (different in length) interlaced with the main tasks. The mean beta power calculated from the EEG signal recorded from the two prefrontal electrodes (Fp1 and Fp2) was used as a stress index. The main results are as follows: (i) we confirmed the previous finding that beta power assessed from the EEG signal recorded from prefrontal electrodes is significantly higher for the STRESS condition compared to NON-STRESS condition; (ii) we found a significant difference in beta power between STRESS conditions that differed in length-the beta power was four times higher for short, compared to long, stress-inducing stimuli; (iii) we did not find enough evidence to confirm (or reject) the hypothesis that stress stimuli repeated over time evoke the habituation effect; although the general trends aggregated over subjects and stressors were negative, their slopes were not statistically significant; moreover, there was no agreement among subjects with respect to the slope of individual trends.

Serotonergic Psychedelics in Neural Plasticity.

Psychedelics, compounds that can induce dramatic changes in conscious experience, have been used by humans for centuries. Recent studies have shown that certain psychedelics can induce neural plasticity by promoting neurite growth and synapse formation. In this review, we focus on the role of classical serotonergic psychedelics in neural plasticity and discuss its implication for their therapeutic potentials.

Can Developments in Tissue Optical Clearing Aid Super-Resolution Microscopy Imaging?

The rapid development of super-resolution microscopy (SRM) techniques opens new avenues to examine cell and tissue details at a nanometer scale. Due to compatibility with specific labelling approaches, in vivo imaging and the relative ease of sample preparation, SRM appears to be a valuable alternative to laborious electron microscopy techniques. SRM, however, is not free from drawbacks, with the rapid quenching of the fluorescence signal, sensitivity to spherical aberrations and light scattering that typically limits imaging depth up to few micrometers being the most pronounced ones. Recently presented and robustly optimized sets of tissue optical clearing (TOC) techniques turn biological specimens transparent, which greatly increases the tissue thickness that is available for imaging without loss of resolution. Hence, SRM and TOC are naturally synergistic techniques, and a proper combination of these might promptly reveal the three-dimensional structure of entire organs with nanometer resolution. As such, an effort to introduce large-scale volumetric SRM has already started; in this review, we discuss TOC approaches that might be favorable during the preparation of SRM samples. Thus, special emphasis is put on TOC methods that enhance the preservation of fluorescence intensity, offer the homogenous distribution of molecular probes, and vastly decrease spherical aberrations. Finally, we review examples of studies in which both SRM and TOC were successfully applied to study biological systems.

Stress induces microglia-associated synaptic circuit alterations in the dorsomedial prefrontal cortex.

The mammalian dorsomedial prefrontal cortex (dmPFC) receives diverse inputs and plays important roles in adaptive behavior and cognitive flexibility. Stress, a major risk factor for many psychiatric disorders, compromises the structure and function of multiple brain regions and circuits. Here we show that 7-day restraint stress impairs reversal learning in the 4-choice odor discrimination test, a decision-making task requiring an intact dmPFC. In vivo two-photon imaging further reveals that stress increases dmPFC dendritic spine elimination, particularly those of the mushroom morphology, without affecting spine formation. In addition, stress alters dmPFC microglial branching complexity and elevates their terminal process dynamics. In stressed mice, dmPFC microglia contact dendrites more frequently, and dendritic spines with microglial contact are prone to elimination. In summary, our work suggests that stress-induced changes in glial-synapse interaction contributes to synaptic loss in dmPFC, resulting in neuronal circuit deficits and impaired cognitive flexibility.

An analog of psychedelics restores functional neural circuits disrupted by unpredictable stress.

Psychological stress affects a wide spectrum of brain functions and poses risks for many mental disorders. However, effective therapeutics to alleviate or revert its deleterious effects are lacking. A recently synthesized psychedelic analog tabernanthalog (TBG) has demonstrated anti-addictive and antidepressant potential. Whether TBG can rescue stress-induced affective, sensory, and cognitive deficits, and how it may achieve such effects by modulating neural circuits, remain unknown. Here we show that in mice exposed to unpredictable mild stress (UMS), administration of a single dose of TBG decreases their anxiety level and rescues deficits in sensory processing as well as in cognitive flexibility. Post-stress TBG treatment promotes the regrowth of excitatory neuron dendritic spines lost during UMS, decreases the baseline neuronal activity, and enhances whisking-modulation of neuronal activity in the somatosensory cortex. Moreover, calcium imaging in head-fixed mice performing a whisker-dependent texture discrimination task shows that novel textures elicit responses from a greater proportion of neurons in the somatosensory cortex than do familiar textures. Such differential response is diminished by UMS and is restored by TBG. Together, our study reveals the effects of UMS on cortical neuronal circuit activity patterns and demonstrate that TBG combats the detrimental effects of stress by modulating basal and stimulus-dependent neural activity in cortical networks.

PSD-95 in CA1 Area Regulates Spatial Choice Depending on Age.

Cognitive processes that require spatial information rely on synaptic plasticity in the dorsal CA1 area (dCA1) of the hippocampus. Since the function of the hippocampus is impaired in aged individuals, it remains unknown how aged animals make spatial choices. Here, we used IntelliCage to study behavioral processes that support spatial choices of aged female mice living in a group. As a proxy of training-induced synaptic plasticity, we analyzed the morphology of dendritic spines and the expression of a synaptic scaffold protein, PSD-95. We observed that spatial choice training in young adult mice induced correlated shrinkage of dendritic spines and downregulation of PSD-95 in dCA1. Moreover, long-term depletion of PSD-95 by shRNA in dCA1 limited correct choices to a reward corner, while reward preference was intact. In contrast, old mice used behavioral strategies characterized by an increased tendency for perseverative visits and social interactions. This strategy resulted in a robust preference for the reward corner during the spatial choice task. Moreover, training decreased the correlation between PSD-95 expression and the size of dendritic spines. Furthermore, PSD-95 depletion did not impair place choice or reward preference in old mice. Thus, our data indicate that while young mice require PSD-95-dependent synaptic plasticity in dCA1 to make correct spatial choices, old animals observe cage mates and stick to a preferred corner to seek the reward. This strategy is resistant to the depletion of PSD-95 in the CA1 area. Overall, our study demonstrates that aged mice combine alternative behavioral and molecular strategies to approach and consume rewards in a complex environment.SIGNIFICANCE STATEMENT It remains poorly understood how aging affects behavioral and molecular processes that support cognitive functions. It is, however, essential to understand these processes to develop therapeutic interventions that support successful cognitive aging. Our data indicate that while young mice require PSD-95-dependent synaptic plasticity in dCA1 to make correct spatial choices (i.e., choices that require spatial information), old animals observe cage mates and stick to a preferred corner to seek the reward. This strategy is resistant to the depletion of PSD-95 in the CA1 area. Overall, our study demonstrates that aged mice combine alternative behavioral and molecular strategies to approach and consume rewards in a complex environment. Second, the contribution of PSD-95-dependent synaptic functions in spatial choice changes with age.

Total Absence of Dystrophin Expression Exacerbates Ectopic Myofiber Calcification and Fibrosis and Alters Macrophage Infiltration Patterns.

Duchenne muscular dystrophy (DMD) causes severe disability and death of young men because of progressive muscle degeneration aggravated by sterile inflammation. DMD is also associated with cognitive and bone-function impairments. This complex phenotype results from the cumulative loss of a spectrum of dystrophin isoforms expressed from the largest human gene. Although there is evidence for the loss of shorter isoforms having impact in the central nervous system, their role in muscle is unclear. We found that at 8 weeks, the active phase of pathology in dystrophic mice, dystrophin-null mice (mdxßgeo) presented with a mildly exacerbated phenotype but without an earlier onset, increased serum creatine kinase levels, or decreased muscle strength. However, at 12 months, mdxßgeo diaphragm strength was lower, whereas fibrosis increased, compared with mdx. The most striking features of the dystrophin-null phenotype were increased ectopic myofiber calcification and altered macrophage infiltration patterns, particularly the close association of macrophages with calcified fibers. Ectopic calcification had the same temporal pattern of presentation and resolution in mdxßgeo and mdx muscles, despite significant intensity differences across muscle groups. Comparison of the rare dystrophin-null patients against those with mutations affecting full-length dystrophins may provide mechanistic insights for developing more effective treatments for DMD.

Whole-body clearing, staining and screening of calcium deposits in the mdx mouse model of Duchenne muscular dystrophy.

BACKGROUND: Duchenne muscular dystrophy (DMD) is a fatal, X-linked genetic disorder. Although DMD is the most common form of muscular dystrophy, only two FDA-approved drugs were developed to delay its progression. In order to assess therapies for treating DMD, several murine models have recently been introduced. As the wide variety of murine models enlighten mechanisms underlying DMD pathology, the question on how to monitor the progression of the disease within the entire musculoskeletal system still remains to be answered. One considerable approach to monitor such progression is histological evaluation of calcium deposits within muscle biopsies. Although accurate, histology is limited to small tissue area and cannot be utilized to evaluate systemic progression of DMD. Therefore, we aimed to develop a methodology suitable for rapid and high-resolution screening of calcium deposits within the entire murine organism. METHODS: Procedures were performed on adult male C57BL/10-mdx and adult male C57BL mice. Animals were sacrificed, perfused, paraformaldehyde-fixed, and subjected to whole-body clearing using optimized perfusion-based CUBIC protocol. Next, cleared organisms were stained with alizarin red S to visualize calcium deposits and subjected to imaging. RESULTS: Study revealed presence of calcium deposits within degenerated muscles of the entire C57BL/10-mdx mouse organism. Calcified deposits were observed within skeletal muscles of the forelimb, diaphragm, lumbar region, pelvic region, and hindlimb. Calcified deposits found in quadriceps femoris, triceps brachii, and spinalis pars lumborum were characterized. Analysis of cumulative frequency distribution showed different distribution characteristics of calcified deposits in quadriceps femoris muscle in comparison to triceps brachii and spinalis pars lumborum muscles (p < 0.001) and quadriceps femoris vs spinalis pars lumborum (p < 0.001). Differences between the number of calcified deposits in selected muscles, their volume, and average volume were statistically significant. CONCLUSIONS: In aggregate, we present new methodology to monitor calcium deposits in situ in the mouse model of Duchenne muscular dystrophy. Sample imaging with the presented setup is feasible and applicable for whole-organ/body imaging. Accompanied by the development of custom-made LSFM apparatus, it allows targeted and precise characterization of calcium deposits in cleared muscles. Hence, presented approach might be broadly utilized to monitor degree to which muscles of the entire organism are affected by the necrosis and how is it altered by the treatment or physical activity of the animal. We believe that this would be a valuable tool for studying organs alternations in a wide group of animal models of muscle dystrophy and bone-oriented diseases.

Autophosphorylation of alpha isoform of calcium/calmodulin-dependent kinase II regulates alcohol addiction-related behaviors.

The development of addiction is associated with a dysregulation of glutamatergic transmission in the brain reward circuit. α isoform of calcium/calmodulin-dependent kinase II (αCaMKII) is one of the key proteins that regulates structural and functional plasticity of glutamatergic synapses. αCaMKII activity can be controlled by the autophosphorylation of threonine 286. The role of this autophosphorylation in the regulation of addiction-related behaviors has been proposed but is still poorly understood. Here, using αCaMKII autophosphorylation-deficient mutant mice (T286A), we show that, in comparison with wild-type animals, they are less resistant to high doses of alcohol and do not show psychostimulant response neither to alcohol injections nor during voluntary alcohol drinking. T286A mutants are also less prone to develop alcohol addiction-related behaviors including an increased motivation for alcohol, persistent alcohol seeking during withdrawal and alcohol consumption on relapse. Finally, we demonstrate that αCaMKII autophosphorylation regulates also alcohol-induced remodeling of glutamatergic synapses in the hippocampus and amygdala. In conclusion, our data suggest that αCaMKII autophosphorylation-dependent remodeling of glutamatergic synapses is a plausible mechanism for the regulation of the alcohol addiction-related behaviors.

Simultaneous Two-photon in Vivo Imaging of Synaptic Inputs and Postsynaptic Targets in the Mouse Retrosplenial Cortex.

This video shows the craniotomy procedure that allows chronic imaging of neurons in the mouse retrosplenial cortex (RSC) using in vivo two-photon microscopy in Thy1-GFP transgenic mouse line. This approach creates a possibility to investigate the correlation of behavioural manipulations with changes in neuronal morphology in vivo. The cranial window implantation procedure was considered to be limited only to the easily accessible cortex regions such as the barrel field. Our approach allows visualization of neurons in the highly vascularized RSC. RSC is an important element of the brain circuit responsible for spatial memory, previously deemed to be problematic for in vivo two-photon imaging. The cranial window implantation over the RSC is combined with an injection of mCherry-expressing recombinant adeno-associated virus (rAAV(mCherry)) into the dorsal hippocampus. The expressed mCherry spreads out to axonal projections from the hippocampus to RSC, enabling the visualization of changes in both presynaptic axonal boutons and postsynaptic dendritic spines in the cortex. This technique allows long-term monitoring of experience-dependent structural plasticity in RSC.

Mapping fear memory consolidation and extinction-specific expression of JunB.

Understanding the molecular and cellular process specifically regulated during fear memory consolidation and extinction is a critical step toward development of new strategies in the treatment of human fear disorders. Here we used inhibitory component of AP-1 transcription factor, JunB, in order to map brain regions where JunB-dependent transcription is regulated during consolidation and extinction of contextual fear memory. We found that contextual fear memory consolidation induced JunB expression in the medial nucleus and intercalated cells of the amygdala while extinction training induced JunB in the CA1 and CA3 areas of the dorsal hippocampus. JunB upregulation induced by contextual fear memory extinction was absent in alphaCaMKII autophosphorylation-deficient mice which have impaired contextual fear memory extinction. Thus, our data suggest that JunB expression in the medial nucleus and intercalated cells of the amygdala is involved in fear memory consolidation while alphaCaMKII-autophosphorylation-dependent JunB expression in the areas CA1 and CA3 of the dorsal hippocampus regulates fear memory extinction.