Exposure to the neurotoxin trimethyltin (TMT) selectively induces hippocampal neuronal injury and astrocyte activation accompanied with resultant neuroinflammation, which causes severe behavioral, cognitive, and memory impairment. A large body of evidence suggests that flaxseed oil (FSO), as one of the richest sources of essential omega-3 fatty acids, i.e., α-linolenic acids (ALA), displays neuroprotective properties. Here, we report the preventive effects of dietary FSO treatment in a rat model of TMT intoxication. The administration of FSO (1 mL/kg, orally) before and over the course of TMT intoxication (a single dose, 8 mg/kg, i.p.) reduced hippocampal cell death, prevented the activation of astrocytes, and inhibited their polarization toward a pro-inflammatory/neurotoxic phenotype. The underlying protective mechanism was delineated through the selective upregulation of BDNF and PI3K/Akt and the suppression of ERK activation in the hippocampus. Pretreatment with FSO reduced cell death and efficiently suppressed the expression of inflammatory molecules. These beneficial effects were accompanied by an increased intrahippocampal content of n-3 fatty acids. In vitro, ALA pretreatment prevented the TMT-induced polarization of cultured astrocytes towards the pro-inflammatory spectrum. Together, these findings support the beneficial neuroprotective properties of FSO/ALA against TMT-induced neurodegeneration and accompanied inflammation and hint at a promising preventive use of FSO in hippocampal degeneration and dysfunction.

We have recently shown delayed increases in GABAB receptor (GABABR) subunit protein levels in the hippocampal dentate gyrus (DG), but not in the pyramidal CA1 and CA3 regions, at 15-30 days after the systemic single administration of trimethyltin (TMT) in mice. An attempt was thus made to determine whether the delayed increases return to the control levels found in naive mice afterward. In the DG on hippocampal slices obtained at 90 days after the administration, however, marked increases were still seen in protein levels of both GABABR1 and GABABR2 subunits without significant changes in calbindin and glial fibrillary acidic protein (GFAP) levels on immunoblotting analysis. Fluoro-Jade B staining clearly revealed the absence of degenerated neurons from the DG at 90 days after the administration. Although co-localization was invariably detected between GABABR2 subunit and GFAP in the DG at 30 days on immunohistochemical analysis, GABABR2-positive cells did not merge well with GFAP-positive cells in the DG at 90 days. These results suggest that both GABABR1 and GABABR2 subunits would be tardily and sustainably up-regulated by cells other than neurons and astrocytes in the murine DG at 90 days after a systemic single injection of TMT.

Reliable fluid biomarkers for evaluating neurotoxicity have yet to be established. However, recent studies have reported neurofilament light chain as a fluid biomarker of several neurodegenerative disorders. In this study, we investigated changes in the cerebrospinal fluid and plasma levels of neurofilament light chain in mice treated with trimethyltin as a neurotoxicant. Trimethyltin diluted with saline was administered by intraperitoneal injection to mice at dose levels of 0 (vehicle control), 1.0, and 2.6 mg/kg body weight (dosage volume: 10 mL/kg). At 3 or 7 days after administration, animals were euthanized by exsanguination under 2-3% isoflurane inhalation anesthesia. Increased neurofilament light chain levels in both the cerebrospinal fluid and plasma were observed in animals from the trimethyltin 2.6 mg/kg body weight group, which indicated the brain lesions including neuronal cell death. Animals from the trimethyltin 1.0 mg/kg body weight group exhibited changes neither in neurofilament light chain levels in the cerebrospinal fluid and plasma nor in the histopathology of the brain at any time point. These data indicate that plasma neurofilament light chain can serve as a useful peripheral biomarker for detecting brain lesions such as neuronal necrosis in mice.

We investigated the neurological effects of the varied erinacine composition of different mycelia cultures in mice with trimethyltin (TMT)-induced neurodegeneration. Forty male ICR mice were randomly divided into five groups of Sham-veh, TMT-veh, TMT-EME, TMT-EMR and TMT-EME/R. The TMT groups received 2.6 mg/kg one-time intraperitoneal injections of TMT. Oral dosages of 200 mg/kg erinacine combination from each Hericium erinaceus mycelia (EM) cultivated formula (100% eucalyptus wood [E], 100% rubber wood [R], or 40% eucalyptus wood/60% rubber wood [E/R]) were given for two weeks. Spatial learning, memory, flexibility, and anxious behaviour were evaluated alongside brain tissues\' oxidative status and histological analyses. Erinacine composition from EME/R exhibited significant positive effects on spatial learning, memory, flexibility, and anxiety (p < 0.05). These findings emerged concurrently with the significant mitigation of hippocampal lipid peroxidation, CA1 hippocampal, cortical neuron, and corpus callosum white matter degeneration (p < 0.05). These neurological benefits were associated with the EME/R composition of erinacine A, C, D, G, H, I, K and R. The best neuroprotective effect against TMT-induced neurodegeneration in mice is offered by the EME/R erinacine composition according to its anti-lipid peroxidation, its nurturing effect on neuronal and white matter, and mitigation of behavioural deficits.

Gastrointestinal symptoms are common health problems found during aging and neurodegenerative diseases. Trimethyltin-induced rat is known as an animal model of hippocampal degeneration with no data on enteric neurodegeneration. This study aimed to investigate the effect of trimethyltin (TMT) induction on the gastrointestinal tract. A 28-day animal study with male Sprague-Dawley rats (3 months old, 150-200 g) given a single TMT injection (8 mg/kg body weight, intraperitoneal) was conducted. The number of neurons in the colonic myenteric plexus was measured using stereological estimation. Histological scoring of colon inflammation, immunohistochemistry of tumor necrosis factor-α (TNF-α), and quantitative PCR were conducted. This study showed neuronal loss in the colonic myenteric plexus of TMT-induced rat model of neurodegeneration. Minor colon inflammation characterized by inflammatory cell infiltration and slightly higher expression of TNF-α in the colon mucosa were observed in the TMT-induced rat. However, the gut microbiota composition of the TMT-induced rat was not different from that of the control rats. This study demonstrates that TMT induces colonic myenteric plexus neurodegeneration and minor colon inflammation, which suggests the potential of this animal model to elucidate the communication between the gastrointestinal tract and central nervous system in neurodegenerative diseases.

BACKGROUND: It has been demonstrated that reactive astrocytes can be polarized into pro-inflammatory A1 phenotype or anti-inflammatory A2 phenotype under neurotoxic and neurodegenerative conditions. Microglia have been suggested to play a critical role in astrocyte phenotype polarization by releasing pro- and anti-inflammatory mediators. In this study, we examined whether trimethyltin (TMT) insult can induce astrocyte polarization in the dentate gyrus of mice, and whether protein kinase Cδ (PKCδ) plays a role in TMT-induced astrocyte phenotype polarization.
METHODS: Male C57BL/6 N mice received TMT (2.6 mg/kg, i.p.), and temporal changes in the mRNA expression of A1 and A2 phenotype markers were evaluated in the hippocampus. In addition, temporal and spatial changes in the protein expression of C3, S100A10, Iba-1, and p-PKCδ were examined in the dentate gyrus. Rottlerin (5 mg/kg, i.p. × 5 at 12-h intervals) was administered 3-5 days after TMT treatment, and the expression of A1 and A2 transcripts, p-PKCδ, Iba-1, C3, S100A10, and C1q was evaluated 6 days after TMT treatment.
RESULTS: TMT treatment significantly increased the mRNA expression of A1 and A2 phenotype markers, and the increased expression of A1 markers remained longer than that of A2 markers. The immunoreactivity of the representative A1 phenotype marker, C3 and A2 phenotype marker, S100A10 peaked 6 days after TMT insult in the dentate gyrus. While C3 was expressed evenly throughout the dentate gyrus, S100A10 was highly expressed in the hilus and inner molecular layer. In addition, TMT insult induced microglial p-PKCδ expression. Treatment with rottlerin, a PKCδ inhibitor, decreased Iba-1 and C3 expression, but did not affect S100A10 expression, suggesting that PKCδ inhibition attenuates microglial activation and A1 astrocyte phenotype polarization. Consistently, rottlerin significantly reduced the expression of C1q and tumor necrosis factor-α (TNFα), which has been suggested to be released by activated microglia and induce A1 astrocyte polarization.
CONCLUSIONS: We demonstrated the temporal and spatial profiles of astrocyte polarization after TMT insult in the dentate gyrus of mice. Taken together, our results suggest that PKCδ plays a role in inducing A1 astrocyte polarization by promoting microglial activation and consequently increasing the expression of pro-inflammatory mediators after TMT insult.

Neurodegeneration implies progressive neuronal loss and neuroinflammation further contributing to pathology progression. It is a feature of many neurological disorders, most common being Alzheimer\'s disease (AD). Repetitive transcranial magnetic stimulation (rTMS) is a non-invasive stimulation which modulates excitability of stimulated brain areas through magnetic pulses. Numerous studies indicated beneficial effect of rTMS in several neurological diseases, including AD, however, exact mechanism are yet to be elucidated. We aimed to evaluate the effect of intermittent theta burst stimulation (iTBS), an rTMS paradigm, on behavioral, neurochemical and molecular level in trimethyltin (TMT)-induced Alzheimer\'s-like disease model. TMT acts as a neurotoxic agent targeting hippocampus causing cognitive impairment and neuroinflammation, replicating behavioral and molecular aspects of AD. Male Wistar rats were divided into four experimental groups-controls, rats subjected to a single dose of TMT (8 mg/kg), TMT rats subjected to iTBS two times per day for 15 days and TMT sham group. After 3 weeks, we examined exploratory behavior and memory, histopathological and changes on molecular level. TMT-treated rats exhibited severe and cognitive deficit. iTBS-treated animals showed improved cognition. iTBS reduced TMT-induced inflammation and increased anti-inflammatory molecules. We examined PI3K/Akt/mTOR signaling pathway which is involved in regulation of apoptosis, cell growth and learning and memory. We found significant downregulation of phosphorylated forms of Akt and mTOR in TMT-intoxicated animals, which were reverted following iTBS stimulation. Application of iTBS produces beneficial effects on cognition in of rats with TMT-induced hippocampal neurodegeneration and that effect could be mediated via PI3K/Akt/mTOR signaling pathway, which could candidate this protocol as a potential therapeutic approach in neurodegenerative diseases such as AD.

Trimethyltin (TMT) is an environmental neurotoxin that mediates dopaminergic neuronal injury in the brain. In this study, we characterized the toxic mechanism and possible protective compounds against TMT-induced neurotoxicity in human dopaminergic neuroblastoma SH-SY5Y cells. Antioxidants such as melatonin, N-acetylcysteine (NAC), α-tocopherol, and allopurinol alleviated TMT toxicity. Apoptosis induced by TMT was identified by altered expression of cleaved caspase-3, Bax, Bcl-2, and Bcl-xL through Western blot analysis. The iron chelator deferoxamine ameliorated the alteration of apoptosis-related proteins through TMT exposure. TMT also induced delayed ultrastructural necrotic features such as mitochondrial swelling and cytoplasmic membrane rupture; NAC reduced these necrotic injuries. Esculetin, meloxicam, celecoxib, and phenidone decreased TMT toxicity. Elevation of the pro-inflammatory cytokines IL-1β, TNF-α, and NF-ĸB and reduction of the antioxidant enzymes catalase and glutathione peroxidase-1 (GPx-1) were induced by TMT and ameliorated by inhibitors of LOX and COX-2 enzymes. Both NMDA and non-NMDA antagonists attenuated TMT toxicity. The free calcium ion modulators nimodipine and BAPTA/AM contributed to neuronal survival against TMT toxicity. Inhibitors of the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin pathway, an autophagy regulator, decreased TMT toxicity. These results imply that TMT neurotoxicity is the chief participant in LOX- and COX-2-mediated apoptosis, partly via necrosis and autophagy in SH-SY5Y cells.

Trimethyltin (TMT) is an irreversible neurotoxicant. Because prenatal TMT exposure has been reported to induce behavioral changes, this study was conducted to observe gender differences and epigenetic changes using a mouse model. In behavioral testing of offspring at 5 weeks of age, the total times spent in the center, corner, or border zones in the male prenatal TMT-exposed mice were less than those of control unexposed mice in the open-field test. Female TMT-exposed mice scored lower on total numbers of arm entries and percentages of alternations than controls in the Y-maze test with lower body weight. We found that only TMT-exposed males had fewer copies of mtDNA in the hippocampus and prefrontal cortex region than controls. Additional epigenetic changes, including increased 5-methyl cytosine/5-hydroxymethyl cytosine levels in the male TMT hippocampus, were observed. After methylation binding domain (MBD) sequencing, multiple signaling pathways related to metabolism and neurodevelopment, including FoxO signaling, were identified by pathway analysis for differentially methylated regions (DMRs). Increased FOXO3 and decreased ASCL1 expression were also observed in male TMT hippocampi. This study suggests that sex differences and epigenetics should be more carefully considered in prenatal toxicology studies.

The organotin, trimethyltin (TMT), is a highly toxic compound. In this study, silver-stained rat brain sections were qualitatively and quantitatively evaluated for degeneration after systemic treatment with TMT. Degenerated neurons were counted using image analysis methods available in the HALO image analysis software. Specific brain areas including the cortex, inferior and superior colliculus, and thalamus were quantitatively analyzed. Our results indicate extensive and widespread damage to the rat brain after systemic administration of TMT. Qualitative results suggest severe TMT-induced toxicity 3 and 7 days after the administration of TMT. Trimethyltin toxicity was greatest in the hippocampus, olfactory area, cerebellum, pons, mammillary nucleus, inferior and superior colliculus, hypoglossal nucleus, thalamus, and cerebellar Purkinje cells. Quantification showed that the optic layer of the superior colliculus exhibited significantly more degeneration compared to layers above and below. The inferior colliculus showed greater degeneration in the dorsal area relative to the central area. Similarly, in cortical layers, there was greater neurodegeneration in deeper layers compared to superficial layers. Quantification of damage in various thalamic nuclei showed that the greatest degeneration occurred in midline and intralaminar nuclei. These results suggest selective neuronal network vulnerability to TMT-related toxicity in the rat brain.