Unveiling the nongenomic actions of thyroid hormones in adult mammalian brain: The legacy of Mary B. Dratman.

A comprehensive review was conducted to compile the contributions of Mary B. Dratman and studies by other researchers in the field of nongenomic actions of thyroid hormones in adult mammalian brain. Dratman and her collaborators authored roughly half of the papers in this area. It has been almost fifty years since Dratman introduced the novel concept of thyroid hormones as neurotransmitters for the first time. The characterization of unique brain-region specific accumulation of thyroid hormones within the nerve terminals in adult mammals was a remarkable contribution by Dratman. It suggested a neurotransmitter- or neuromodulator-like role of thyroid hormone and/or its derivative, 3-iodothyronamine within adrenergic systems in adult mammalian brain. Several studies by other researchers using synaptosomes as a model system, have contributed to the concept of direct nongenomic actions of thyroid hormones at synaptic regions by establishing that thyroid hormones or their derivatives can bind to synaptosomal membranes, alter membrane functions including enzymatic activities and ion transport, elicit Ca2+/NO-dependent signaling pathways and induce substrate-protein phosphorylation. Such findings can help to explain the physiological and pathophysiological roles of thyroid hormone in psychobehavioral control in adult mammalian brain. However, the exact mode of nongenomic actions of thyroid hormones at nerve terminals in adult mammalian brain awaits further study.

Integrated network-based multiple computational analyses for identification of co-expressed candidate genes associated with neurological manifestations of COVID-19.

'Tripartite network' (TN) and 'combined gene network' (CGN) were constructed and their hub-bottleneck and driver nodes (44 genes) were evaluated as 'target genes' (TG) to identify 21 'candidate genes' (CG) and their relationship with neurological manifestations of COVID-19. TN was developed using neurological symptoms of COVID-19 found in literature. Under query genes (TG of TN), co-expressed genes were identified using pair-wise mutual information to genes available in RNA-Seq autopsy data of frontal cortex of COVID-19 victims. CGN was constructed with genes selected from TN and co-expressed in COVID-19. TG and their connecting genes of respective networks underwent functional analyses through findings of their enrichment terms and pair-wise 'semantic similarity scores' (SSS). A new integrated 'weighted harmonic mean score' was formulated assimilating values of SSS and STRING-based 'combined score' of the selected TG-pairs, which provided CG-pairs with properties of CGs as co-expressed and 'indispensable nodes' in CGN. Finally, six pairs sharing seven 'prevalent CGs' (ADAM10, ADAM17, AKT1, CTNNB1, ESR1, PIK3CA, FGFR1) showed linkages with the phenotypes (a) directly under neurodegeneration, neurodevelopmental diseases, tumour/cancer and cellular signalling, and (b) indirectly through other CGs under behavioural/cognitive and motor dysfunctions. The pathophysiology of 'prevalent CGs' has been discussed to interpret neurological phenotypes of COVID-19.

Matrix metallopeptidase 9 as a host protein target of chloroquine and melatonin for immunoregulation in COVID-19: A network-based meta-analysis.

AIMS: The molecular pathogenesis of COVID-19 is similar to other coronavirus (CoV) infections viz. severe acute respiratory syndrome (SARS) in human. Due to scarcity of the suitable treatment strategy, the present study was undertaken to explore host protein(s) targeted by potent repurposed drug(s) in COVID-19. MATERIALS AND METHODS: The differentially expressed genes (DEGs) were identified from microarray data repository of SARS-CoV patient blood. The repurposed drugs for COVID-19 were selected from available literature. Using DEGs and drugs, the protein-protein interaction (PPI) and chemo-protein interaction (CPI) networks were constructed and combined to develop an interactome model of PPI-CPI network. The top-ranked sub-network with its hub-bottleneck nodes were evaluated with their functional annotations. KEY FINDINGS: A total of 120 DEGs and 65 drugs were identified. The PPI-CPI network (118 nodes and 293 edges) exhibited a top-ranked sub-network (35 nodes and 174 connectivities) with 12 hub-bottleneck nodes having two drugs chloroquine and melatonin in association with 10 proteins corresponding to six upregulated and four downregulated genes. Two drugs interacted directly with the hub-bottleneck node i.e. matrix metallopeptidase 9 (MMP9), a host protein corresponding to its upregulated gene. MMP9 showed functional annotations associated with neutrophil mediated immunoinflammation. Moreover, literature survey revealed that angiotensin converting enzyme 2, a membrane receptor of SARS-CoV-2 virus, might have functional cooperativity with MMP9 and a possible interaction with both drugs. SIGNIFICANCE: The present study reveals that between chloroquine and melatonin, melatonin appears to be more promising repurposed drug against MMP9 for better immunocompromisation in COVID-19.

Alterations of thyroidal status in brain regions and hypothalamo-pituitary-blood-thyroid-axis associated with dopaminergic depletion in substantia nigra and ROS formation in different brain regions after MPTP treatment in adult male mice.

MPTP produces oxidative stress, damages niagrostriatal dopaminergic neurons and develops Parkinsonism in rodents. Due to paucity of information, the thyroidal status in brain regions and peripheral tissues during different post-treatment days in MPTP-induced mice had been executed in the present study. MPTP depleted tyrosine hydroxylase protein expressions that signify the dopaminergic neuronal damage in substantia nigra. MPTP elevated ROS formation differentially in brain regions (cerebral cortex, hippocampus, substantia nigra) with maximal elevation at hippocampus. The changes in thyroid hormone (T4 and T3) levels indicate that brain regions might combat the adverse situation by keeping the levels of thyroid hormones either unchanged or in the elevated conditions in the latter phases (day-3 and day-7), apart from the depletion of thyroid hormones in certain brain regions (T4 in SN and hippocampus, T3 in hippocampus) as the immediate (day-1) effects after MPTP treatment. MPTP caused alterations of cellular morphology, RNA:Protein ratio and TPO protein expression, concomitantly depleted TPO mRNA expression and elevated TSH levels in the thyroid gland. Although T4 levels changed differentially, T3 levels remained unaltered in thyroid gland throughout the post-treatment days. Results have been discussed mentioning the putative role of T4 and TSH in apoptosis and/or proliferation/differentiation of thyrocytes. In blood, T4 levels remained unchanged while the changes in T3 and TSH levels did not signify the clinical feature of hypo/hyperthyroidism of animals. In the pituitary, both T4 and T3 levels remained elevated where TSH differentially altered (elevated followed by depletion) during post-treatment days. Notably, T4, T3 and TSH levels did not alter in hypothalamus except initial (day-1) depletion of the T4 level. Therefore, the feedback control mechanism of hypothalamo-pituitary-blood-thyroid-axis failed to occur after MPTP treatment. Overall, MPTP altered thyroidal status in the brain and peripheral tissues while both events might occur in isolation as well.

Sex and estrous cycle dependent changes in locomotor activity, anxiety and memory performance in aged mice after exposure of light at night.

Light-at-night (LAN) can affect mammalian behaviour. But, the effects of LAN on aged rodents remain undefined yet. In the present investigation, aged Swiss Albino mice, habituated in regular light-dark cycle, were exposed to bright-light-pulse (1-h) at night on the day of study followed by experimentations for assessment of locomotor activities in the open field, anxiety in the elevated plus maze and short-term memory for novel object recognition (NOR) in the habituated field. Under without-bright-light exposure, (a) aged proestrous females showed greater locomotor activities and less anxiety than in aged diestrous females, (b) aged males showed locomotor activities and anxiety level similar to aged diestrous females and aged proestrous females respectively and (c) all animals failed to retain in object discrimination memory. LAN exposure exhibited the continual failure of such retention of memory while animals showed free and spontaneous exploration with thigmotactic behaviour having no object bias and/or phobia, but time stay in objects by animals altered variably among sexes and stages of estrous cycle. Overall, the LAN caused (a) diminution in locomotor activities, rise in anxiety and failure of memory for recognition of both familiar and novel objects in aged proestrous females, (b) hyperlocomotor activities and reduction in anxiety in both males and diestrous females with the failure of memory for recognition of novel objects only in aged males while diestrous females showed enhanced exploration time to both objects during NOR. Thus, nocturnal behaviour of aged mice varies with sex and estrous cycle and light acts differentially on them.

Age related rise in lactate and its correlation with lactate dehydrogenase (LDH) status in post-mitochondrial fractions isolated from different regions of brain in mice.

Rise in brain lactate is the hallmark of ageing. Separate studies report that ageing is associated with elevation of lactate level and alterations of lactate dehydrogenase (LDH)-A/B mRNA-expression-ratio in cerebral cortex and hippocampus. However, age related lactate rise in brain and its association with LDH status and their brain regional variations are still elusive. In the present study, level of lactate, LDH (A and B) activity and LDH-A expression were evaluated in post-mitochondrial fraction of tissues isolated from four different brain regions (cerebral cortex, hippocampus, substantia nigra and cerebellum) of young and aged mice. Lactate levels elevated in four brain regions with maximum rise in substantia nigra of aged mice. LDH-A protein expression and its activity decreased in cerebral cortex, hippocampus and substantia nigra without any changes of these parameters in cerebellum of aged mice. LDH-B activity decreased in hippocampus, substantia nigra and cerebellum whereas its activity remains unaltered in cerebral cortex of aged mice. Accordingly, the ratio of LDH-A/LDH-B-activity remains unaltered in hippocampus and substantia nigra, decreased in cerebral cortex and increased in cerebellum. Therefore, rise of lactate in three brain regions (cerebral cortex, hippocampus, substantia nigra) appeared to be not correlated with the alterations of its regulatory enzymes activities in these three brain regions, rather it supports the fact of involvement of other mechanisms, like lactate transport and/or aerobic/anaerobic metabolism as the possible cause(s) of lactate rise in these three brain regions. The increase in LDH-A/LDH-B-activity-ratio appeared to be positively correlated with elevated lactate level in cerebellum of aged mice. Overall, the present study indicates that the mechanism of rise in lactate in brain varies with brain regions where LDH status plays an important role during ageing.

TNFR2 mediated TNF-α signaling and NF-κB activation in hippocampus of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated mice.

1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine (MPTP) -induced neuroinflammation and its impact in hippocampus remain elusive till date. Our present study includes the time dependent changes of inflammatory molecules in mouse hippocampus during MPTP treatment. MPTP treatment increased level of TNF-α, enhanced expression of TNFR2 along with PI3 kinase (PI3K) induced phosphorylation of Akt resulting in persistent nuclear factor-κB (NF-κB) activation. The expressions gradually increased from Day1 post-MPTP treatment, maximally at Day3 post-treatment. MPTP induced translocation of p65 and p52, two subunits of NF-κB family, to nucleus where they had been found to dimerize. Therefore, MPTP induced TNF-α signaling through TNFR2 mediated pathway and recruited p65-p52 dimer in hippocampal nucleus which is reported to have protective effect on hippocampal neurons indicated by unchanged neuronal count in hippocampus in treated groups with respect to control. Our finding suggests that this unique NF-κB dimer plays some role in providing inherent protection to hippocampus during MPTP-treatment.

Gender features and estrous cycle variations of nocturnal behavior of mice after a single exposure to light at night.

Light at night alters behavior and cognitive performances in rodents, the variations of which in gender and stages of reproductive cycle in females are elusive. Young mice habituated in light:dark (12:12h) cycle were given a single exposure of light (100lx) at early night for one hour duration followed by experimentations in open field (closed wall with circular big arena), elevated plus maze and square habituated field for memory performance using novel object recognition task. Light effects were compared with results found during without light conditions. Proestrous females appeared to have greater locomotor activity, less anxiety and better memory performance compared to the diestrous females at night without light exposure. The status of locomotor activity, anxiety and memory performance of male mice at night without light exposure appeared to be comparable to females where the stage of estrous cycle is important to characterize the nocturnal behavior of male mice. Light maximally affected proestrous females with decrease in locomotor activity, increase in anxiety and failure of memory performance. Male and diestrous female mice performed memory performance without alteration of locomotor activity and anxiety after exposure to light where males performed better memory performance with greater locomotor activity and more anxiety compared to that of diestrous females. The present study characterizes the mice nocturnal behavior with and without a single exposure to light stimuli with its gender features and estrous cycle variation. In addition, the study indicates an association of memory performance with locomotor activity and anxiety in mice nocturnal behavior.

Alteration of nuclear factor-kappaB pathway promote neuroinflammation depending on the functions of estrogen receptors in substantia nigra after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine treatment.

The simultaneous role of neuroprotective estrogen and neurodegenerative inflammation during the progression of Parkinson's disease (PD) is still remaining elusive. The novel importance of the present study in MPTP mediated mouse model of Parkinson's disease (PD) is-to investigate the status of neuronal and glial cells in a time chase experiment; to explore which pathway of NF-kappaB exist to proceed the neuroinflammation; to investigate the status of estrogen and the activation pattern of nuclear or cytosolic estrogen receptors in either sexes of Swiss albino mice during MPTP mediated progressive neurodegeneration in the substantia nigra. After MPTP intoxication, the nigral molecular anatomy was changed differently in separate time interval during the progression of neurodegeneration with/without association of glial cells and functional (via its nuclear and cytosolic receptors) estrogen level. Both the canonical and/or non-canonical pathways of NF-kappaB exist in the substantia nigra of both the sexes after MPTP treatment that is why inspite of presence of estrogen, neuroinflammation progresses. The homodimeric or heterodimeric form of ER-beta binds with NF-kappaB molecules p65 and RelB differently, but the canonical or non-canonical pathways of NF-kappaB molecules could not be stopped or may be promoted.

Alteration in Nuclear Factor-KappaB Pathway and Functionality of Estrogen via Receptors Promote Neuroinflammation in Frontal Cortex after 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Treatment.

The MPTP mediated neurodegeneration in substantia nigra has been well studied, but not the status of frontal cortex. The novelty of the present study is to explore the sex difference of frontal cortex during MPTP intoxication and to investigate the role of estrogen and its receptors in presence of glial cells in a time chase experiment; to identify which pathway of NF-kappaB exist to proceed the neuroinflammation; to investigate the estrogen binding with its nuclear or cytosolic receptors and whether any direct relation exists between estrogen receptor (ER) -beta and NF-kappaB molecules p65 and RelB. The progression of neurodegeneration occurred with the association of glial cells and functional (via its nuclear and cytosolic receptors) estrogen level. Both the canonical and/or non canonical pathways of NF-kappaB exist in frontal cortex of both the sexes after MPTP treatment. The homodimeric or heterodimeric form of ER-beta binds with NF-kappaB molecules p65 and RelB differently, but the canonical or non canonical pathways of NF-kappaB molecules could not be stopped or may be promoted. The changes in the molecular and cellular pattern in frontal cortex of both sexes during MPTP intoxication depends on the estrogen function via its nuclear or cytosolic estrogen receptors.

MRI characterization of temporal lobe epilepsy using rapidly measurable spatial indices with hemisphere asymmetries and gender features.

INTRODUCTION: The paucity of morphometric markers for hemispheric asymmetries and gender variations in hippocampi and amygdalae in temporal lobe epilepsy (TLE) calls for better characterization of TLE by finding more useful prognostic MRI parameter(s). METHODS: T1-weighted MRI (3 T) morphometry using multiple parameters of hippocampus-parahippocampus (angular and linear measures, volumetry) and amygdalae (volumetry) including their hemispheric asymmetry indices (AI) were evaluated in both genders. The cutoff values of parameters were statistically estimated from measurements of healthy subjects to characterize TLE (57 patients, 55% male) alterations. RESULTS: TLE had differential categories with hippocampal atrophy, parahippocampal angle (PHA) acuteness, and several other parametric changes. Bilateral TLE categories were much more prevalent compared to unilateral TLE categories. Female patients were considerably more disposed to bilateral TLE categories than male patients. Male patients displayed diverse categories of unilateral abnormalities. Few patients (both genders) had combined bilateral appearances of hippocampal atrophy, amygdala atrophy, PHA acuteness, and increase in hippocampal angle (HA) where medial distance ratio (MDR) varied among genders. TLE had gender-specific and hemispheric dominant alterations in AI of parameters. Maximum magnitude of parametric changes in TLE includes (a) AI increase in HA of both genders, (b) HA increase (bilateral) in female patients, and (c) increase in ratio of amygdale/hippocampal volume (unilateral, right hemispheric), and AI decrease in MDR, in male patients. CONCLUSION: Multiparametric MRI studies of hippocampus and amygdalae, including their hemispheric asymmetry, underscore better characterization of TLE. Rapidly measurable single-slice parameters (HA, PHA, MDR) can readily delineate TLE in a time-constrained clinical setting, which contrasts with customary three-dimensional hippocampal volumetry that requires many slice computation.

Differential regional expression patterns of α-synuclein, TNF-α, and IL-1ß; and variable status of dopaminergic neurotoxicity in mouse brain after Paraquat treatment.

BACKGROUND: Paraquat (1, 1-dimethyl-4, 4-bipyridium dichloride; PQ) causes neurotoxicity, especially dopaminergic neurotoxicity, and is a supposed risk factor for Parkinson's disease (PD). However, the cellular and molecular mechanisms of PQ-induced neurodegeneration are far from clear. Previous studies have shown that PQ induces neuroinflammation and dopaminergic cell loss, but the prime cause of those events is still in debate. METHODS: We examined the neuropathological effects of PQ not only in substantia nigra (SN) but also in frontal cortex (FC) and hippocampus of the progressive mouse (adult Swiss albino) model of PD-like neurodegeneration, using immunohistochemistry, western blots, and histological and biochemical analyses. RESULTS: PQ caused differential patterns of changes in cellular morphology and expression of proteins related to PD and neuroinflammation in the three regions examined (SN, FC and hippocampus). Coincident with behavioral impairment and brain-specific ROS generation, there was differential immunolocalization and decreased expression levels of tyrosine hydroxylase (TH) in the three regions, whereas α-synuclein immunopositivity increased in hippocampus, increased in FC and decreased in SN. PQ-induced neuroinflammation was characterized by area-specific changes in localization and appearances of microglial cells with or without activation and increment in expression patterns of tumor necrosis factor-α in the three regions of mouse brain. Expression of interleukin-1ß was increased in FC and hippocampus but not significantly changed in SN. CONCLUSION: The present study demonstrates that PQ induces ROS production and differential α-synuclein expression that promotes neuroinflammation in microglia-dependent or -independent manners, and produces different patterns of dopaminergic neurotoxicity in three different regions of mouse brain.

Expression of mRNA and proteins for GnRH I and II and their receptors in primate corpus luteum during menstrual cycle.

The differential expression of mRNA and protein of GnRH I, II and their receptors (RI and RII) in the monkey corpus luteum (CL) were measured during different stages of the luteal phase of the menstrual cycle as an initial step towards considering the role and regulation of GnRH (I and II) system during luteinization and luteolysis in primates. RT-PCR confirmed the sequence identity of PCR products and real time PCR quantified specific mRNA expressions. Proteins were localized by immunohistochemistry (IHC). Changes in mRNA expression patterns of GnRH I and II (increased) and GnRH RII (decreased) were maximal at mid-late to late stages, that is, at CL regression, where as GnRH RI was low during the entire luteal phase. However, RT-PCR and IHC studies confirmed the presence of GnRH RI at both mRNA and protein levels, respectively. IHC results showed the presence of GnRH I, II and their receptors in steroidogenic cells (granulose-luteal cells and thecal-luteal cells) across the luteal phase. Hence, GnRH I and II systems may have a role on both luteinization (from early to mid stages of CL) and luteolysis (from mid-late to very-late stages of CL). These novel findings suggest that monkey luteal GnRH system may have a role in fertility regulation in paracrine and/or autocrine manner.

Presence of immunoreactive gonadotropin releasing hormone (GnRH) and its receptor (GnRHR) in rat ovary during pregnancy.

The present study aims at quantification of gonadotropin releasing hormone (GnRH) by radioimmunoassay, relative expression of its mRNA by real-time PCR accompanied by its cellular localization in the rat ovary by immunonohistochemistry (IHC) during different time points of pregnancy. To determine the involvement of endogenous ovarian GnRH in receptor mediated local autocrine/paracrine functions within the ovary, the cell specific localization of the classical receptor for GnRH (GnRHR) in the ovary by IHC and expression pattern of its mRNA were studied during pregnancy. Receptor expression during each time point within the ovary was reconfirmed by Western blot analysis accompanied by densitometric analysis of the signal intensity. Results reveal that the content of ovarian GnRH reaches its maximum on Day 20. The densitometric analysis of GnRHR receptor expression from Western blot study exhibits a decreasing trend by Day 20. Presence of GnRH and GnRHR mRNA in the ovary indicates the local synthesis of both ligand and receptor in the rat ovary. Differential expression of GnRH/GnRHR in the corpus luteum throughout pregnancy strengthens the hypothesis of the involvement of ovarian GnRH in local ovarian functions by receptor-mediated mechanisms. The expression of GnRH and GnRHR in the atretic antral follicles is indicative of the possible involvement of this decapeptide in processes like follicular atresia. The expression of GnRH/GnRHR in the nonatretic antral follicles and their oocytes requires further in-depth investigation. Collectively, this study for the first time reveals the presence of endogenous ovarian GnRH/GnRHR supporting their possible involvement in local autocrine/paracrine functions during pregnancy.

Localization of immunoreactive gonadotropin-releasing hormone and relative expression of its mRNA in the oviduct during pregnancy in rats.

This study was designed to determine the cellular and ultrastructural distribution of the gonadotropin-releasing hormone (GnRH) and the relative expression of its mRNA in the oviduct of rats during different time points (days 7, 9, 16, and 20) of pregnancy. Immunofluorescent localization and confocal microscopic techniques were used to determine the cellular distribution of GnRH in the oviduct. Immunogold electron microscopy indicated its localization at the ultrastructural level, and real-time PCR was used to study the expression pattern of GnRH mRNA in the oviduct during pregnancy. In general, GnRH was localized within the epithelial cells lining the oviductal lumen at each selected time point. A strong correlation between the fluorescence intensity of GnRH-immunoreactive cells and the relative expression of GnRH mRNA was noted on days 7 and 16, followed by a plateau by day 20. At the ultrastructural level, uniform labeling of colloidal gold particles was observed in secretory vesicles and lamella of the luminal epithelium as well as the lumen of the oviduct. Collectively, these results demonstrate for the first time that the oviductal epithelium synthesizes and secretes the decapeptide GnRH during pregnancy in rats, which may have a possible role in postimplantation embryonic development and the maintenance of pregnancy.

Stimulation of AChE activity in relation to changes in electronmicroscopic structure of adult rat cerebrocortical synaptosomes pretreated with 3-5-3'-triiodo-L-thyronine.

Triiodothyronine (T3) stimulated AChE activity in depolarization-induced intact synaptosomes (isolated from adult rat cerebral cortex) suspended in calcium-supplemented choline chloride buffer in a time-dependent manner maximally 45-60 s after T3 administration and in a dose-dependent manner with an optimum at 10-100 nM. T3 (100 nM) had no such effects on AChE activity in synaptosomes at non-depolarized conditions. There was no direct effect of T3 on AChE activity of lysed synaptosomal suspension in the physiological range (nM) of T3. The experiments suggest that T3 might have a role in the calcium-dependent release/co-release of acetylcholine from intact synaptosomes concomitant with the acceleration of choline uptake mechanisms that has been reported to accompany elevation of AChE activity. Additionally, electron microscopic structures showed condensation of the cytosolic content with increase in electron density, formation of intrasynaptosomal coarse vesicles and appearance of vesicular fusion like structures (meandering) at the periphery in depolarization-induced T3-treated (60 s) intact synaptosomes, indicating the occurrence of the release of neurotransmitters. The present investigation indicates a definite role of T3 on Ca2+-dependent cholinergic neurotransmission.