Light affects behavioral despair involving the clock gene Period 1.

Light at night has strong effects on physiology and behavior of mammals. It affects mood in humans, which is exploited as light therapy, and has been shown to reset the circadian clock in the suprachiasmatic nuclei (SCN). This resetting is paramount to align physiological and biochemical timing to the environmental light-dark cycle. Here we provide evidence that light at zeitgeber time (ZT) 22 affects mood-related behaviors also in mice by activating the clock gene Period1 (Per1) in the lateral habenula (LHb), a brain region known to modulate mood-related behaviors. We show that complete deletion of Per1 in mice led to depressive-like behavior and loss of the beneficial effects of light on this behavior. In contrast, specific deletion of Per1 in the region of the LHb did not affect mood-related behavior, but suppressed the beneficial effects of light. RNA sequence analysis in the mesolimbic dopaminergic system revealed profound changes of gene expression after a light pulse at ZT22. In the nucleus accumbens (NAc), sensory perception of smell and G-protein coupled receptor signaling were affected the most. Interestingly, most of these genes were not affected in Per1 knock-out animals, indicating that induction of Per1 by light serves as a filter for light-mediated gene expression in the brain. Taken together we show that light affects mood-related behavior in mice at least in part via induction of Per1 in the LHb with consequences on mood-related behavior and signaling mechanisms in the mesolimbic dopaminergic system.

Airway inflammation and dysbiosis in antibody deficiency despite the presence of IgG.

BACKGROUND: Patients with antibody deficiency suffer chronic respiratory symptoms, recurrent exacerbations, and progressive airways disease despite systemic replacement of IgG. Little is known about the respiratory tract biology of these patients. OBJECTIVE: We sought to measure immunoglobulin levels, inflammatory cytokines, and mediators of tissue damage in serum and sputum from patients with antibody deficiency and healthy controls; to analyze the respiratory microbiome in the same cohorts. METHODS: We obtained paired sputum and serum samples from 31 immunocompetent subjects and 67 antibody-deficient patients, the latter divided on computed tomography scan appearance into "abnormal airways" (bronchiectasis or airway thickening) or "normal airways." We measured inflammatory cytokines, immunoglobulin levels, neutrophil elastase, matrix-metalloproteinase-9, urea, albumin, and total protein levels using standard assays. We used V3-V4 region 16S sequencing for microbiome analysis. RESULTS: Immunodeficient patients had markedly reduced IgA in sputum but higher concentrations of IgG compared with healthy controls. Inflammatory cytokines and tissue damage markers were higher in immunodeficient patients, who also exhibited dysbiosis with overrepresentation of pathogenic taxa and significantly reduced alpha diversity compared with immunocompetent individuals. These differences were seen regardless of airway morphology. Sputum matrix-metalloproteinase-9 and elastase correlated inversely with alpha diversity in the antibody-deficient group, as did sputum IgG, which correlated positively with several inflammatory markers, even after correction for albumin levels. CONCLUSIONS: Patients with antibody deficiency, even with normal lung imaging, exhibit inflammation and dysbiosis in their airways despite higher levels of IgG compared with healthy controls.

A novel soluble complement receptor 1 fragment with enhanced therapeutic potential.

Human complement receptor 1 (HuCR1) is a pivotal regulator of complement activity, acting on all three complement pathways as a membrane-bound receptor of C3b/C4b, C3/C5 convertase decay accelerator, and cofactor for factor I-mediated cleavage of C3b and C4b. In this study, we sought to identify a minimal soluble fragment of HuCR1, which retains the complement regulatory activity of the wildtype protein. To this end, we generated recombinant, soluble, and truncated versions of HuCR1 and compared their ability to inhibit complement activation in vitro using multiple assays. A soluble form of HuCR1, truncated at amino acid 1392 and designated CSL040, was found to be a more potent inhibitor than all other truncation variants tested. CSL040 retained its affinity to both C3b and C4b as well as its cleavage and decay acceleration activity and was found to be stable under a range of buffer conditions. Pharmacokinetic studies in mice demonstrated that the level of sialylation is a major determinant of CSL040 clearance in vivo. CSL040 also showed an improved pharmacokinetic profile compared with the full extracellular domain of HuCR1. The in vivo effects of CSL040 on acute complement-mediated kidney damage were tested in an attenuated passive antiglomerular basement membrane antibody-induced glomerulonephritis model. In this model, CSL040 at 20 and 60 mg/kg significantly attenuated kidney damage at 24 h, with significant reductions in cellular infiltrates and urine albumin, consistent with protection from kidney damage. CSL040 thus represents a potential therapeutic candidate for the treatment of complement-mediated disorders.

Lack of drug-induced post-retrieval amnesia for auditory fear memories in rats.

BACKGROUND: Long-term memory formation is generally assumed to involve the permanent storage of recently acquired memories, making them relatively insensitive to disruption, a process referred to as memory consolidation. However, when retrieved under specific circumstances, consolidated fear memories are thought to return to a labile state, thereby opening a window for modification (e.g., attenuation) of the memory. Several interventions during a critical time frame after this destabilization seem to be able to alter the retrieved memory, for example by pharmacologically interfering with the restabilization process, either by direct protein synthesis inhibition or indirectly, using drugs that can be safely administered in patients (e.g., propranolol). Here, we find that, contrary to expectations, systemic pharmacological manipulations in auditory fear-conditioned rats do not lead to drug-induced post-retrieval amnesia. RESULTS: In a series of well-powered auditory fear conditioning experiments (four with propranolol, 10 mg/kg, two with rapamycin, 20-40 mg/kg, one with anisomycin, 150 mg/kg and cycloheximide, 1.5 mg/kg), we found no evidence for reduced cued fear memory expression during a drug-free test in adult male Sprague-Dawley rats that had previously received a systemic drug injection upon retrieval of the tone fear memory. All experiments used standard fear conditioning and reactivation procedures with freezing as the behavioral read-out (conceptual or exact replications of published reports) and common pharmacological agents. Additional tests confirmed that the applied drug doses and administration routes were effective in inducing their conventional effects on expression of fear (propranolol, acutely), body weight (rapamycin, anisomycin, cycloheximide), and consolidation of extinction memories (cycloheximide). CONCLUSIONS: In contrast with previously published studies, we did not find evidence for drug-induced post-retrieval amnesia, underlining that this effect, as well as its clinical applicability, may be considerably more constrained and less readily reproduced than what the current literature would suggest.

Limited replicability of drug-induced amnesia after contextual fear memory retrieval in rats.

With the ultimate goal of investigating boundary conditions for post-reactivation amnesia, we set out to replicate studies in which systemic, post-reactivation administration of midazolam, propranolol, or cycloheximide resulted in amnesia for contextual fear memories. Our experiments involved conceptual as well as exact replications of previously published studies. In most of our experiments, we adopted a procedure that conformed to the standard 3-day protocol typically used in the literature, with contextual fear conditioning on day 1, unreinforced re-exposure to the conditioning context followed by systemic injection of the amnestic drug on day 2, and a memory retention test on day 3. Given the plethora of successful studies with large effects sizes and the absence of any failed replications in the literature, we were surprised to find that we were generally unable to replicate those findings. Our results suggest that post-reactivation amnesia by systemic drug administration in rats is more difficult to obtain than what would be expected based on published empirical reports. At present, it remains unclear which conditions determine the success of this procedure.

Face categorization and behavioral templates in rats.

Rodents have become a popular model in vision science. It is still unclear how vision in rodents relates to primate vision when it comes to complex visual tasks. Here we report on the results of training rats in a face-categorization and generalization task. Additionally, the Bubbles paradigm is used to determine the behavioral templates of the animals. We found that rats are capable of face categorization and can generalize to previously unseen exemplars. Performance is affected-but remains above chance-by stimulus modifications such as upside-down and contrast-inverted stimuli. The behavioral templates of the rats overlap with a pixel-based template, with a bias toward the upper left parts of the stimuli. Together, these findings significantly expand the evidence about the extent to which rats learn complex visual-categorization tasks.

No harmful effect of propranolol administered prior to fear memory extinction in rats and humans.

Exposure therapy is an evidence-based treatment option for anxiety-related disorders. Many patients also take medication that could, in principle, affect exposure therapy efficacy. Clinical and laboratory evidence indeed suggests that benzodiazepines may have detrimental effects. Large clinical trials with propranolol, a common beta-blocker, are currently lacking, but several preclinical studies do indicate impaired establishment of safety memories. Here, we investigated the effects of propranolol given prior to extinction training in 9 rat studies (N = 215) and one human study (N = 72). A Bayesian meta-analysis of our rat studies provided strong evidence against propranolol-induced extinction memory impairment during a drug-free test, and the human study found no significant difference with placebo. Two of the rat studies actually suggested a small beneficial effect of propranolol. Lastly, two rat studies with a benzodiazepine (midazolam) group provided some evidence for a harmful effect on extinction memory, i.e., impaired extinction retention. In conclusion, our midazolam findings are in line with prior literature (i.e., an extinction retention impairment), but this is not the case for the 10 studies with propranolol. Our data thus support caution regarding the use of benzodiazepines during exposure therapy, but argue against a harmful effect of propranolol on extinction learning.

The importance of contrast features in rat vision.

Models of object recognition have mostly focused upon the hierarchical processing of objects from local edges up to more complex shape features. An alternative strategy that might be involved in pattern recognition centres around coarse-level contrast features. In humans and monkeys, the use of such features is most documented in the domain of face perception. Given prior suggestions that, generally, rodents might rely upon contrast features for object recognition, we hypothesized that they would pick up the typical contrast features relevant for face detection. We trained rats in a face-nonface categorization task with stimuli previously used in computer vision and tested for generalization with new, unseen stimuli by including manipulations of the presence and strength of a range of contrast features previously identified to be relevant for face detection. Although overall generalization performance was low, it was significantly modulated by contrast features. A model taking into account the summed strength of contrast features predicted the variation in accuracy across stimuli. Finally, with deep neural networks, we further investigated and quantified the performance and representations of the animals. The findings suggest that rat behaviour in visual pattern recognition tasks is partially explained by contrast feature processing.

A computationally informed comparison between the strategies of rodents and humans in visual object recognition.

Many species are able to recognize objects, but it has been proven difficult to pinpoint and compare how different species solve this task. Recent research suggested to combine computational and animal modelling in order to obtain a more systematic understanding of task complexity and compare strategies between species. In this study, we created a large multidimensional stimulus set and designed a visual discrimination task partially based upon modelling with a convolutional deep neural network (CNN). Experiments included rats (N = 11; 1115 daily sessions in total for all rats together) and humans (N = 45). Each species was able to master the task and generalize to a variety of new images. Nevertheless, rats and humans showed very little convergence in terms of which object pairs were associated with high and low performance, suggesting the use of different strategies. There was an interaction between species and whether stimulus pairs favoured early or late processing in a CNN. A direct comparison with CNN representations and visual feature analyses revealed that rat performance was best captured by late convolutional layers and partially by visual features such as brightness and pixel-level similarity, while human performance related more to the higher-up fully connected layers. These findings highlight the additional value of using a computational approach for the design of object recognition tasks. Overall, this computationally informed investigation of object recognition behaviour reveals a strong discrepancy in strategies between rodent and human vision.

Mice lacking circadian clock components display different mood-related behaviors and do not respond uniformly to chronic lithium treatment.

Genomic studies suggest an association of circadian clock genes with bipolar disorder (BD) and lithium response in humans. Therefore, we tested mice mutant in various clock genes before and after lithium treatment in the forced swim test (FST), a rodent behavioral test used for evaluation of depressive-like states. We find that expression of circadian clock components, including Per2, Cry1 and Rev-erbα, is affected by lithium treatment, and thus, these clock components may contribute to the beneficial effects of lithium therapy. In particular, we observed that Cry1 is important at specific times of the day to transmit lithium-mediated effects. Interestingly, the pathways involving Per2 and Cry1, which regulate the behavior in the FST and the response to lithium, are distinct as evidenced by the phosphorylation of GSK3ß after lithium treatment and the modulation of dopamine levels in the striatum. Furthermore, we observed the co-existence of depressive and mania-like symptoms in Cry1 knock-out mice, which resembles the so-called mixed state seen in BD patients. Taken together our results strengthen the concept that a defective circadian timing system may impact directly or indirectly on mood-related behaviors.

Rhythm and mood: relationships between the circadian clock and mood-related behavior.

Mood disorders are multifactorial and heterogeneous diseases caused by the interplay of several genetic and environmental factors. In humans, mood disorders are often accompanied by abnormalities in the organization of the circadian system, which normally synchronizes activities and functions of cells and tissues. Studies on animal models suggest that the basic circadian clock mechanism, which runs in essentially all cells, is implicated in the modulation of biological phenomena regulating affective behaviors. In particular, recent findings highlight the importance of the circadian clock mechanisms in neurological pathways involved in mood, such as monoaminergic neurotransmission, hypothalamus-pituitary-adrenal axis regulation, suprachiasmatic nucleus and olfactory bulb activities, and neurogenesis. Defects at the level of both, the circadian clock mechanism and system, may contribute to the etiology of mood disorders. Modification of the circadian system using chronotherapy appears to be an effective treatment for mood disorders. Additionally, understanding the role of circadian clock mechanisms, which affect the regulation of different mood pathways, will open up the possibility for targeted pharmacological treatments.

The nuclear receptor REV-ERBα regulates Fabp7 and modulates adult hippocampal neurogenesis.

The function of the nuclear receptor Rev-erbα (Nr1d1) in the brain is, apart from its role in the circadian clock mechanism, unknown. Therefore, we compared gene expression profiles in the brain between wild-type and Rev-erbα knock-out (KO) animals. We identified fatty acid binding protein 7 (Fabp7, Blbp) as a direct target of repression by REV-ERBα. Loss of Rev-erbα manifested in memory and mood related behavioral phenotypes and led to overexpression of Fabp7 in various brain areas including the subgranular zone (SGZ) of the hippocampus, where neuronal progenitor cells (NPCs) can initiate adult neurogenesis. We found increased proliferation of hippocampal neurons and loss of its diurnal pattern in Rev-erbα KO mice. In vitro, proliferation and migration of glioblastoma cells were affected by manipulating either Fabp7 expression or REV-ERBα activity. These results suggest an important role of Rev-erbα and Fabp7 in adult neurogenesis, which may open new avenues for treatment of gliomas as well as neurological diseases such as depression and Alzheimer.

Role of the circadian clock gene Per2 in adaptation to cold temperature.

Adaptive thermogenesis allows mammals to resist to cold. For instance, in brown adipose tissue (BAT) the facultative uncoupling of the proton gradient from ATP synthesis in mitochondria is used to generate systemic heat. However, this system necessitates an increase of the Uncoupling protein 1 (Ucp1) and its activation by free fatty acids. Here we show that mice without functional Period2 (Per2) were cold sensitive because their adaptive thermogenesis system was less efficient. Upon cold-exposure, Heat shock factor 1 (HSF1) induced Per2 in the BAT. Subsequently, PER2 as a co-activator of PPARα increased expression of Ucp1. PER2 also increased Fatty acid binding protein 3 (Fabp3), a protein important to transport free fatty acids from the plasma to mitochondria to activate UCP1. Hence, in BAT PER2 is important for the coordination of the molecular response of mice exposed to cold by synchronizing UCP1 expression and its activation.

Protein phosphatase 1 (PP1) is a post-translational regulator of the mammalian circadian clock.

Circadian clocks coordinate the timing of important biological processes. Interconnected transcriptional and post-translational feedback loops based on a set of clock genes generate and maintain these rhythms with a period of about 24 hours. Many clock proteins undergo circadian cycles of post-translational modifications. Among these modifications, protein phosphorylation plays an important role in regulating activity, stability and intracellular localization of clock components. Several protein kinases were characterized as regulators of the circadian clock. However, the function of protein phosphatases, which balance phosphorylation events, in the mammalian clock mechanism is less well understood. Here, we identify protein phosphatase 1 (PP1) as regulator of period and light-induced resetting of the mammalian circadian clock. Down-regulation of PP1 activity in cells by RNA interference and in vivo by expression of a specific inhibitor in the brain of mice tended to lengthen circadian period. Moreover, reduction of PP1 activity in the brain altered light-mediated clock resetting behavior in mice, enhancing the phase shifts in either direction. At the molecular level, diminished PP1 activity increased nuclear accumulation of the clock component PER2 in neurons. Hence, PP1, may reduce PER2 phosphorylation thereby influencing nuclear localization of this protein. This may at least partially influence period and phase shifting properties of the mammalian circadian clock.