Datumetine Preferentially Upregulates N-methyl-D-aspartate Receptor Signalling Pathways in Different Brain Regions of Mice.

Introduction: We previously reported that datumetine possesses binding affinity with N-methyl-D-aspartate receptor (NMDAR) and that 14-day exposure to datumetine altered NMDAR signaling by mimicking glutamate toxicity. Here, we investigated the potential neuroprotective effect of a single shot of a low dose of datumetine administration in BALB/c mice. Methods: 30 male adult BALB/c mice were used for the study. The mice were randomly divided into three groups of ten mice each with an intraperitoneal injection of 0.1 mL of 10% DMSO for the Vehicle group, Datumetine group were administered 0.1 mg/kg body weight (bw) of datumetine and MK-801+Datumetine group were administered 0.5 mg/kg bw of MK-801 (to block NMDAR) followed by 0.1 mg/kg bw of datumetine after 30 minutes. 24 hours after administration, mice were euthanized in an isoflurane chamber followed by perfusion with 1X PBS. Brains were excised and stored at -20°C till further processing. Mice designated for IHC were further perfused with 4% PFA and brain excised and stored in 4% PFA till further processing. NMDAR signalling molecules expression was evaluated in frozen brain samples and the fixed brain samples were stained for neuron, vGlut and NMDAR subtypes. Results: Relative to vehicle (Veh), datumetine downregulate calcium calmodulin kinase II alpha (CamKIIα) expression in the hippocampus and prefrontal cortex (PFC) but not in the cerebellum, cyclic AMP response element binding protein (CREB) was also upregulated only in the PFC but phosphorylated CREB (pCREB) was also upregulated in three brain regions observed, while brain-derived neurotrophic factor (BDNF) was only upregulated in hippocampus and PFC of Datumetine relative to vehicle (Veh). On the other hand, dizocilpine (MK-801) reversed some of the effects of datumetine in the observed brain regions. No major histological alterations were observed in the different brain regions immunohistochemically. Conclusion: We conclude that a low dose of datumetine moderately enhances NMDAR activity. This showed the neuroprotective potentials of low datumetine exposure.

Oral thymoquinone modulates cyclophosphamide-induced testicular toxicity in adolescent Wistar rats.

Cyclophosphamide (CYP) is an effective anti-cancer drug that is widely accepted, but it is not devoid of unintended toxic effects. Gonadal toxicity is reported as one of the side effects of its long-time use. This study examined the effects of thymoquinone (TQ) on the biological integrities of the testes after cyclophosphamide exposure. Thirty adolescent male Wistar rats (100-110 g) were divided into six groups (n = 5), receiving normal saline (NS), 20 mg/kg of CYP (CYP), 5 mg/kg of TQ (TQ5), 10 mg/kg of TQ (TQ10), 20 mg/kg of CYP and 5 mg/kg of TQ (CTQ5), and 20 mg/kg of CYP and 10 mg/kg of TQ (CTQ10) respectively. On the 22nd day, blood, semen and testicular samples were collected for the assay of serum reproductive hormones (follicle-stimulating (FSH) and luteinizing (LH) hormones), semen analysis and testicular histology and proliferating cell nuclear antigen (PCNA) expression. The results revealed that CYP exposure affected functional and structural integrities of the testes, by depleting sperm count and motility, testosterone, LH, spermatogenic and mature sperm cell population, Leydig cells and PCNA immunoreactive proliferating cells. TQ interventions were able to reverse all cytotoxic CYP impacts, but with differential activities on the hormonal concentrations, specifically LH and FSH. Cumulatively, thymoquinone may be a potent agent against cyclophosphamide effects on the physiological, regeneration and histological integrities of the testes, as observed in this study.

Datumetine exposure alters hippocampal neurotransmitters system in C57BL/6 mice.

Our previous study showed that datumetine modulates NMDAR activity with long term exposure leading to memory deficit and altered NMDAR signaling. We aim to explore the neurotransmitters perturbations of acute datumetine-NMDAR interaction. Fifteen C57/BL6 mice were used for the study, they are divided into three groups of 5 animals each. Animals were administered DMSO (DMSO/Control), 0.25 mg/kg body weight of datumetine (0.25 Datumetine) and 1 mg/kg bodyweight of datumetine (1.0 Datumetine) intraperitoneally for 14 days. At the end of treatment, animals were euthanized in isofluorane chamber, perfused transcardially with 1XPBS followed by PFA. Immunofluorescence procedure was done to check the distribution of neurons, astrocytes, microglia and major neuronal subtypes in the hippocampus. Expansion and electron microscopy techniques were used to assess the condition of the synapses. Quantitative data were expressed as mean ± SEM and analyzed using ANOVA with Tukey post hoc using p < 0.05 as significant. Datumetine increased the expression of CD11b, GFAP, vGlut1, GABA, CHRNA7 and TH while expression of TrPH and NeuN were reduced in the hippocampus compared to control animals. Synaptic loss was evident in datumetine exposed animals with reduced synaptic vesicles accompanied by a thickness of postsynaptic density than that of control animals. This study concludes that acute datumetine exposure alters hippocampal neurotransmitter systems.

Effects of datumetine on hippocampal NMDAR activity.

The usage (abuse) of Datura metel is becoming increasingly worrisome among the Nigerian populace especially among the youth considering its side effects such as hallucination. This work was designed to identify the phytochemicals in datura plant that potentially interact with NMDAR as it affects the electrical and memory activities of the brain. Ligand-protein interaction was assessed using autodock vina to identify phytochemicals that can interact with NMDAR. Datumetine was found to have the best interaction fit with NMDAR at both allosteric and orthosteric binding sites. Furthermore, using electrophysiological, behavioural and western blotting techniques, it was observed that the administration of datumetine positively modulates the NMDAR current by prolonging burst duration and interspike interval, induces seizures in C57BL/6 mice. Acute exposure leads to memory deficit on NOR and Y-maze test while immunoblotting results showed increased expression of GluN1 and CamKIIα while pCamKIIα-T286, CREB and BDNF were downregulated. The results showed that the memory deficit seen in datura intoxication is possibly the effects of datumetine on NMDAR.

Chlorpyrifos- and Dichlorvos-Induced Oxidative and Neurogenic Damage Elicits Neuro-Cognitive Deficits and Increases Anxiety-Like Behavior in Wild-Type Rats.

The execution of agricultural activities on an industrial scale has led to indiscriminate deposition of toxic xenobiotics, including organophosphates, in the biome. This has led to intoxication characterized by deleterious oxidative and neuronal changes. This study investigated the consequences of oxidative and neurogenic disruptions that follow exposure to a combination of two organophosphates, chlorpyrifos (CPF) and dichlorvos (DDVP), on neuro-cognitive performance and anxiety-like behaviors in rats. Thirty-two adult male Wistar rats (150⁻170 g) were randomly divided into four groups, orally exposed to normal saline (NS), DDVP (8.8 mg/kg), CPF (14.9 mg/kg), and DDVP + CPF for 14 consecutive days. On day 10 of exposure, anxiety-like behavior and amygdala-dependent fear learning were assessed using open field and elevated plus maze paradigms, respectively, while spatial working memory was assessed on day 14 in the Morris water maze paradigm, following three training trials on days 11, 12, and 13. On day 15, the rats were euthanized, and their brains excised, with the hippocampus and amygdala removed. Five of these samples were homogenized and centrifuged to analyze nitric oxide (NO) metabolites, total reactive oxygen species (ROS), and acetylcholinesterase (AChE) activity, and the other three were processed for histology (cresyl violet stain) and proliferative markers (Ki67 immunohistochemistry). Marked (p ≤0.05) loss in body weight, AChE depletion, and overproduction of both NO and ROS were observed after repeated exposure to individual and combined doses of CPF and DDVP. Insults from DDVP exposure appeared more severe owing to the observed greater losses in the body weights of exposed rats. There was also a significant (p ≤0.05) effect on the cognitive behaviors recorded from the exposed rats, and these deficits were related to the oxidative damage and neurogenic cell loss in the hippocampus and the amygdala of the exposed rats. Taken together, these results provided an insight that oxidative and neurogenic damage are central to the severity of neuro-cognitive dysfunction and increased anxiety-like behaviors that follow organophosphate poisoning.

Histology and ultrastructure of transitional changes in skin morphology in the juvenile and adult four-striped mouse (Rhabdomys pumilio).

The four-striped mouse has a grey to brown coloured coat with four characteristic dark stripes interspersed with three lighter stripes running along its back. The histological differences in the skin of the juvenile and adult mouse were investigated by Haematoxylin and Eosin and Masson Trichrome staining, while melanocytes in the skin were studied through melanin-specific Ferro-ferricyanide staining. The ultrastructure of the juvenile skin, hair follicles, and melanocytes was also explored. In both the juvenile and adult four-striped mouse, pigment-containing cells were observed in the dermis and were homogeneously dispersed throughout this layer. Apart from these cells, the histology of the skin of the adult four-striped mouse was similar to normal mammalian skin. In the juvenile four-striped mouse, abundant hair follicles of varying sizes were observed in the dermis and hypodermis, while hair follicles of similar size were only present in the dermis of adult four-striped mouse. Ultrastructural analysis of juvenile hair follicles revealed that the arrangement and differentiation of cellular layers were typical of a mammal. This study therefore provides unique transition pattern in the four-striped mouse skin morphology different from the textbook description of the normal mammalian skin.