Improving stroke outcomes in hyperglycemic mice by modulating tPA/NMDAR signaling to reduce inflammation and hemorrhages.

ABSTRACT: The pharmacological intervention for ischemic stroke hinges on intravenous administration of the recombinant tissue-type plasminogen activator (rtPA, Alteplase/Actilyse) either as a standalone treatment or in conjunction with thrombectomy. However, despite its clinical significance, broader use of rtPA is constrained because of the risk of hemorrhagic transformations (HTs). Furthermore, the presence of diabetes or chronic hyperglycemia is associated with an elevated risk of HT subsequent to thrombolysis. This detrimental impact of tPA on the neurovascular unit in patients with hyperglycemia has been ascribed to its capacity to induce endothelial N-methyl-D-aspartate receptor (NMDAR) signaling, contributing to compromised blood-brain barrier integrity and neuroinflammatory processes. In a mouse model of thromboembolic stroke with chronic hyperglycemia, we assessed the effectiveness of rtPA and N-acetylcysteine (NAC) as thrombolytic agents. We also tested the effect of blocking tPA/NMDAR signaling using a monoclonal antibody, Glunomab. Magnetic resonance imaging, speckle contrast imaging, flow cytometry, and behavioral tasks were used to evaluate stroke outcomes. In hyperglycemic animals, treatment with rtPA resulted in lower recanalization rates and increased HTs. Conversely, NAC treatment reduced lesion sizes while mitigating HTs. After a single administration, either in standalone or combined with rtPA-induced thrombolysis, Glunomab reduced brain lesion volumes, HTs, and neuroinflammation after stroke, translating into improved neurological outcomes. Additionally, we demonstrated the therapeutic efficacy of Glunomab in combination with NAC or as a standalone strategy in chronic hyperglycemic animals. Counteracting tPA-dependent endothelial NMDAR signaling limits ischemic damages induced by both endogenous and exogenous tPA, including HTs and inflammatory processes after ischemic stroke in hyperglycemic animals.

The Anxiolytic-like Properties of a Tryptic Hydrolysate of Bovine αs1 Casein Containing α-Casozepine Rely on GABAA Receptor Benzodiazepine Binding Sites but Not the Vagus Nerve.

(1) Background: A tryptic hydrolysate of bovine αs1-casein (CH) exerts anxiolytic-like properties in many species, including humans. This is mainly related to the presence of α-casozepine (α-CZP), which yields these properties in rodents. This study evaluates, in a rat model, the roles of the vagus nerve and the benzodiazepine binding site of GABAA receptors in the mode of action of CH. (2) Methods: The conditioned defensive burying test was used to evaluate anxiety. (3) Results: Participation of the vagus nerve in the mode of action of CH was excluded, as the global anxiety score in vagotomised rats was not significantly different from that of non-vagotomised animals. The blocking of the binding sites of benzodiazepines with flumazenil antagonised CH anxiolytic-like properties. (4) Conclusions: The vagus nerve does not play a role in the anxiolytic-like properties of CH. On the other hand, this anxiolytic-like activity relies on the benzodiazepine binding site of the GABAA receptors. This result is consistent with previous in vitro studies and, more specifically with the discovery of α-CZP, the peptide responsible for the anxiolytic-like properties of CH.

Post-acute delivery of α5-GABAA antagonist, S 44819, improves functional recovery in juvenile rats following stroke.

Hypo-excitability was reported in the peri-infarct tissue following stroke, an effect counteracted by a blockage of α5-GABAA receptors in adult rodents. Our present study aims to evaluate the effect of a selective α5-GABAA receptor antagonist, S 44819, in stroke in juvenile animals. We have set up and characterized an original model of transient ischemic stroke in 28 day-old Sprague-Dawley rats (45-min occlusion of the middle cerebral artery by intraluminal suture). In this model, S 44819 (1, 3 and 10 mg/kg, b.i.d) was orally administered from day 3 to day 16 after stroke onset. Sensorimotor recovery was assessed on day 1, day 9 and day 16 after stroke onset. Results show that rats treated with S 44819 at the doses of 3 and 10 mg/kg displayed a significant improvement of the neurological deficits (neuroscore) on day 9 and day 16, when compared with animals treated with vehicle. Grip-test data analysis reveals that rats treated with S 44819 at the dose of 3 mg/kg displayed a better recovery on day 9 and day 16. These results are in agreement with those previously observed in adult rats, demonstrating that targeting α5-GABAA receptors improves neurological recovery after stroke in juvenile rats.

Evaluation of the antidepressant- and anxiolytic-like effects of a hydrophilic extract from the green seaweed Ulva sp. in rats.

OBJECTIVES: The green seaweed Ulva sp. contains a large amount of ulvans, a family of sulphated polysaccharides. The present study was designed to investigate in rats the antidepressant- and anxiolytic-like effects of a hydrophilic extract of Ulva sp. (MSP) containing about 45% of ulvans. METHODS: After a 14-day administration of MSP at doses of 10, 20 and 40 mg/kg/day, 48 and 60 male adult Wistar rats were respectively tested in the elevated plus-maze (EPM) and the forced swimming test (FST). In the FST, MSP effects were compared to the reference antidepressant drug imipramine (IMI) (10 mg/kg/day). Acute and sub-chronic toxicities of the extract were also assessed in male and female rats following OECD guidelines. RESULTS: MSP treatment did not modify anxiety-related behaviour in the EPM. In contrast, MSP induced a dose-dependent reduction of immobility behaviour in the FST. At the highest tested dose of 40 mg/kg, MSP displayed a significant antidepressant-like effect similar to IMI. MSP did not modify the exploratory behaviour of rats in the open field test and did not produce any toxic effect. DISCUSSION: MSP may potentially represent a good adjunct or alternative to existing antidepressant therapeutics. Further studies are necessary to confirm the mechanism of action of MSP and its modulation of brain functioning.

Benefits of Preventive Administration of Chlorella sp. on Visceral Pain and Cystitis Induced by a Single Administration of Cyclophosphamide in Female Wistar Rat.

Chlorella sp. is a green microalgae containing nutrients, vitamins, minerals, and chlorophyll. In some communities, Chlorella sp. is a traditional medicinal plant used for the management of inflammation-related diseases. In a rat model, ROQUETTE Chlorella sp. (RCs) benefits were investigated on visceral pain and associated inflammatory parameters related to cystitis both induced by cyclophosphamide (CYP). RCs was orally administered every day from day 1-16 (250 and 500 mg/kg body weight). Six hours after an intraperitoneal injection of 200 mg/kg body weight of CYP, body temperature, general behavior, food intake, and body weight were recorded. Twenty-four hours after CYP injection, rats were tested in two behavioral tests, an open field and the aversive light stimulus avoidance conditioning test, to evaluate the influence of pain on general activity and learning ability of rats. After euthanasia, bladders were weighed, their thickness was scored, and the urinary hemoglobin was measured. RCs orally administered at the two dosages significantly reduced visceral pain and associated inflammatory parameters related to cystitis both induced by CYP injection, and improved rat behavior. To conclude, RCs demonstrated beneficial effects against visceral pain and cystitis.

In Situ Microparticles Loaded with S-Nitrosoglutathione Protect from Stroke.

Treatment of stroke, especially during the first hours or days, is still lacking. S-nitrosoglutathione (GSNO), a cerebroprotective agent with short life time, may help if administered early with a sustain delivery while avoiding intensive reduction in blood pressure. We developed in situ forming implants (biocompatible biodegradable copolymer) and microparticles (same polymer and solvent emulsified with an external oily phase) of GSNO to lengthen its effects and allow cerebroprotection after a single subcutaneous administration to Wistar rats. Arterial pressure was recorded for 3 days (telemetry, n = 14), whole-blood platelet aggregation up to 13 days (aggregometry, n = 58), and neurological score, cerebral infarct size and edema volume for 2 days after obstruction of the middle cerebral artery by autologous blood clots (n = 30). GSNO-loaded formulations (30 mg/kg) induced a slighter and longer hypotension (-10 vs. -56 ± 6 mmHg mean arterial pressure, 18 h vs. 40 min) than free GSNO at the same dose. The change in pulse pressure (-50%) lasted even up to 42 h for microparticles. GSNO-loaded formulations (30 mg/kg) prevented the transient 24 h hyper-aggregability observed with free GSNO and 7.5 mg/kg-loaded formulations. When injected 2 h after stroke, GSNO-loaded microparticles (30 mg/kg) reduced neurological score at 24 (-62%) and 48 h (-75%) vs. empty microparticles and free GSNO 7.5 mg/kg and, compared to free GSNO, divided infarct size by 10 and edema volume by 8 at 48 h. Corresponding implants reduced infarct size and edema volume by 2.5 to 3 times. The longer (at least 2 days) but slight effects on arterial pressures show sustained delivery of GSNO-loaded formulations (30 mg/kg), which prevent transient platelet hyper-responsiveness and afford cerebroprotection against the consequences of stroke. In conclusion, in situ GSNO-loaded formulations are promising candidates for the treatment of stroke.

Oral and Topical Administration of ROQUETTE Schizochytrium sp. Alleviate Skin Inflammation and Improve Wound Healing in Mice.

The human body is constantly exposed to the risk of traumatic lesions. ROQUETTE Schizochytrium sp. (SCs) is a marine microalgae containing large amounts of health-valuable nutrients, more particularly polyunsaturated fatty acids such as docosahexaenoic acid. SCs was investigated by oral administration (125, 250 and 500 mg/kg) and cutaneous application (2.5, 5.0 and 10.0%) to evaluate its impact in two dermatological disorder models in mice: skin inflammation and wound healing. For skin inflammation, it was administered during 14 days starting one week before the induction of chronic skin inflammation by repeated cutaneous application of 12-O-tetradecanoylphorbol 13-acetate (TPA). For wound healing the microalgae was administered after incisional wound healing of the skin until complete wound healing. Results indicated that oral and topical administrations of the two higher doses of SCs had significant effects on macroscopic score of skin inflammation. It had also efficient effect on healing process and duration of wound healing with a dose-response by oral administration and a maximal effect observed from the lowest to the highest dose by topical application. These findings suggest that administration of SCs by both oral and topical routes appeared to have beneficial effects on skin lesions.

Mining the brain metabolome to understand behavioural disruptions induced in mouse fed Hypochoeris radicata (L.), a neurotoxic plant for horse.

Mining the brain metabolome to understand behavioural disruptions induced in mouse fed Hypochoeris radicata (L.), a neurotoxic plant for horse. C57BL/6J mice orally exposed to 9% H. radicata (HR) are metabolically competent laboratory animals which can be used as model of Australian stringhalt, a neurological horse disease induced by HR ingestion. So, the present study was conducted to assess the brain metabolome and the behavioural performances of mice fed with a 9%-HR-based diet for 21 days. By the end of the period of exposure, mice were investigated for motor activity and coordination, anxiety level, learning and memory performances, social behaviour and rewarding properties of for the plant. Thus, the animals were sacrificed and the brain metabolome was studied using (1)H NMR spectroscopy. HR-exposed mice displayed a motor hyperactivity in several tasks, a less resignation in the forced swimming test, and paradigm place preference for the plant. A bootstrap-based regularized canonical analysis performed on merged behavioural and metabolic datasets showed a clear relationship in HR-treated mice between an increase in cerebral scyllo-inositol, an increased motor activity, and seemingly rewarding properties of HR. These results underlie the interest of such a dual approach to characterize functional end-points of a pathophysiological model of the Australian stringhalt in equine species.

Assessment of psychotropic-like properties of a probiotic formulation (Lactobacillus helveticus R0052 and Bifidobacterium longum R0175) in rats and human subjects.

In a previous clinical study, a probiotic formulation (PF) consisting of Lactobacillus helveticus R0052 and Bifidobacterium longum R0175 (PF) decreased stress-induced gastrointestinal discomfort. Emerging evidence of a role for gut microbiota on central nervous system functions therefore suggests that oral intake of probiotics may have beneficial consequences on mood and psychological distress. The aim of the present study was to investigate the anxiolytic-like activity of PF in rats, and its possible effects on anxiety, depression, stress and coping strategies in healthy human volunteers. In the preclinical study, rats were daily administered PF for 2 weeks and subsequently tested in the conditioned defensive burying test, a screening model for anti-anxiety agents. In the clinical trial, volunteers participated in a double-blind, placebo-controlled, randomised parallel group study with PF administered for 30 d and assessed with the Hopkins Symptom Checklist (HSCL-90), the Hospital Anxiety and Depression Scale (HADS), the Perceived Stress Scale, the Coping Checklist (CCL) and 24 h urinary free cortisol (UFC). Daily subchronic administration of PF significantly reduced anxiety-like behaviour in rats (P < 0·05) and alleviated psychological distress in volunteers, as measured particularly by the HSCL-90 scale (global severity index, P < 0·05; somatisation, P < 0·05; depression, P < 0·05; and anger-hostility, P < 0·05), the HADS (HADS global score, P < 0·05; and HADS-anxiety, P < 0·06), and by the CCL (problem solving, P < 0·05) and the UFC level (P < 0·05). L. helveticus R0052 and B. longum R0175 taken in combination display anxiolytic-like activity in rats and beneficial psychological effects in healthy human volunteers.

Variations in illumination, closed wall transparency and/or extramaze space influence both baseline anxiety and response to diazepam in the rat elevated plus-maze.

Numerous methodological-related variables have been demonstrated to influence the baseline anxiety level of rodents exposed to the elevated plus-maze (EPM), raising questions about the sensitivity of this test for the detection of the effects of anxiolytic drugs. Thus, the present study was designed (1) to assess the combined effects of illumination (40-lx red or white light), closed wall type (walls made of translucent or opaque material) and extramaze space size (small or spacious experimental room) on rat behaviour, and (2) to investigate the effects of such parameters on the relevance of the maze for detecting the effects of diazepam orally administrated at the anxiolytic dose of 3 mg/kg. Results indicate that illumination and closed wall type are two main independent parameters that are able to modify the open arm avoidance. Moreover, the closed wall type interacts with the extramaze space size since the reduction of the open arm exploration induced by opaque closed walls is two-fold stronger in the spacious experimental room than in the small one. Finally, the diazepam anxiolytic activity is significantly detected in our laboratory in specific EPM conditions (maze with opaque walls, use of a red light, maze located in a spacious experimental room). In conclusion, the present study demonstrates that an inappropriate baseline anxiety level due to the methodological use of the EPM can dramatically reduce the sensitivity of the maze for the detection of benzodiazepine-related compounds. This study also provides new insights into the perception of the EPM open space in rats.

Ethological comparison of the effects of a bovine alpha s1-casein tryptic hydrolysate and diazepam on the behaviour of rats in two models of anxiety.

A bovine alpha s1-casein tryptic hydrolysate was previously demonstrated to display an anxiolytic-like activity in the conditioned defensive burying and in the elevated plus-maze models when i.p. injected. The present study assessed the anxiolytic-like effects of this tryptic hydrolysate after an oral administration in rats faced to the same behavioural situations using diazepam as a reference. In a first experiment, the behavioural effects of the hydrolysate in the conditioned defensive burying test were investigated at doses ranging 5-50 mg/kg. The results showed that the minimal dose required to elicit an anxiolytic-like activity is 15 mg/kg. In a second experiment, the alpha s1-casein tryptic hydrolysate (15 mg/kg, p.o.) was demonstrated to display an anxiolytic-like activity similar to diazepam (3 mg/kg, p.o.) in the conditioned defensive burying test and the elevated plus-maze. However, the ethological analysis of behaviour indicated that this hydrolysate has a different activity compared to diazepam. While diazepam induced a disinhibition state in rats, possibly related to the risk-taking behaviour observed after a benzodiazepine ingestion in humans, the tryptic hydrolysate did not display such a side effect. These results suggest that the mechanism of action of the bovine alpha s1-casein tryptic hydrolysate may differ from that of diazepam.