Induced expression of rabies glycoprotein in the dorsal hippocampus enhances hippocampal dependent memory in a rat model of Alzheimer's disease.

The Rabies virus is a neurotropic virus that manipulates the natural cell death processes of its host to ensure its own survival and replication. Studies have shown that the anti-apoptotic effect of the virus is mediated by one of its protein named, rabies glycoprotein (RVG). Alzheimer's disease (AD) is characterized by the loss of neural cells and memory impairment. We aim to examine whether expression of RVG in the hippocampal cells can shield the detrimental effects induced by Aß. Oligomeric form of Aß (oAß) or vehicle was bilaterally microinjected into the dorsal hippocampus of male Wistar rats. One week later, two µl (108 T.U. /ml) of the lentiviral vector carrying RVG gene was injected into their dorsal hippocampus (post-treatment). In another experiment, the lentiviral vector was microinjected one week before Aß injection (pre-treatment). One week later, the rat's brain was sliced into cross-sections, and the presence of RVG-expressing neuronal cells was confirmed using fluorescent microscopy. Rats were subjected to assessments of spatial learning and memory as well as passive avoidance using the Morris water maze (MWM) and the Shuttle box apparatuses, respectively. Protein expression of AMPA receptor subunit (GluA1) was determined using western blotting technique. In MWM, Aß treated rats showed decelerated acquisition of the task and impairment of reference memory. RVG expression in the hippocampus prevented and restored the deficits in both pre- and post- treatment conditions, respectively. It also improved inhibitory memory in the oAß treated rats. RVG increased the expression level of GluA1 level in the hippocampus. Based on our findings, the expression of RVG in the hippocampus has the potential to enhance both inhibitory and spatial learning abilities, ultimately improving memory performance in an AD rat model. This beneficial effect is likely attributed, at least in part, to the increased expression of GluA1-containing AMPA receptors.

Neuroprotective Effects of Ferrostatin and Necrostatin Against Entorhinal Amyloidopathy-Induced Electrophysiological Alterations Mediated by voltage-gated Ca2+ Channels in the Dentate Gyrus Granular Cells.

Alzheimer's disease (AD) is a progressive neurodegenerative disease that is the main form of dementia. Abnormal deposition of amyloid-beta (Aß) peptides in neurons and synapses cause neuronal loss and cognitive deficits. We have previously reported that ferroptosis and necroptosis were implicated in Aß25-35 neurotoxicity, and their specific inhibitors had attenuating effects on cognitive impairment induced by Aß25-35 neurotoxicity. Here, we aimed to examine the impact of ferroptosis and necroptosis inhibition following the Aß25-35 neurotoxicity on the neuronal excitability of dentate gyrus (DG) and the possible involvement of voltage-gated Ca2+ channels in their effects. After inducing Aß25-35 neurotoxicity, electrophysiological alterations in the intrinsic properties and excitability were recorded by the whole-cell patch-clamp under current-clamp condition. Voltage-clamp recordings were also performed to shed light on the involvement of calcium channel currents. Aß25-35 neurotoxicity induced a considerable reduction in input resistance (Rin), accompanied by a profoundly decreased excitability and a reduction in the amplitude of voltage-gated calcium channel currents in the DG granule cells. However, three days of administration of either ferrostatin-1 (Fer-1), a ferroptosis inhibitor, or Necrostatin-1 (Nec-1), a necroptosis inhibitor, in the entorhinal cortex could almost preserve the normal excitability and the Ca2+ currents. In conclusion, these findings suggest that ferroptosis and necroptosis involvement in EC amyloidopathy could be a potential candidate to prevent the suppressive effect of Aß on the Ca2+ channel current and neuronal function, which might take place in neurons during the development of AD.

Lentiviral Expression of Rabies Virus Glycoprotein in the Rat Hippocampus Strengthens Synaptic Plasticity.

Rabies virus (RABV) is a neurotropic virus exclusively infecting neurons in the central nervous system. RABV encodes five proteins. Among them, the viral glycoprotein (RVG) plays a key role in viral entry into neurons and rabies pathogenesis. It was shown that the nature of the C-terminus of the RABV G protein, which possesses a PDZ-binding motif (PBM), modulates the virulence of the RABV strain. The neuronal protein partners recruited by this PBM may alter host cell function. This study was conducted to investigate the effect of RVG on synaptic function in the hippocampal dentate gyrus (DG) of rat. Two µl (108 T.U./ml) of the lentiviral vector containing RVG gene was injected into the DG of rat hippocampus. After 2 weeks, the rat's brain was cross-sectioned and RVG-expressing cells were detected by fluorescent microscopy. Hippocampal synaptic activity of the infected rats was then examined by recording the local field potentials from DG after stimulation of the perforant pathway. Short-term synaptic plasticity was also assessed by double pulse stimulation. Expression of RVG in DG increased long-term potentiation population spikes (LTP-PS), whereas no facilitation of LTP-PS was found in neurons expressing δRVG (deleted PBM). Furthermore, RVG and δRVG strengthened paired-pulse facilitation. Heterosynaptic long-term depression (LTD) in the DG was significantly blocked in RVG-expressing group compared to the control group. This blockade was dependent to PBM motif as rats expressing δRVG in the DG-expressed LTD comparable to the RVG group. Our data demonstrate that RVG expression facilitates both short- and long-term synaptic plasticity in the DG indicating that it may involve both pre- and postsynaptic mechanisms to alter synaptic function. Further studies are needed to elucidate the underlying mechanisms.

Rabies virus glycoprotein enhances spatial memory via the PDZ binding motif.

Rabies is a life-threatening viral infection of the brain. Rabies virus (RABV) merely infects excitable cells including neurons provoking drastic behaviors including negative emotional memories. RABV glycoprotein (RVG) plays a critical role in RABV pathogenesis. RVG interacts with various cytoplasmic PDZ (PSD-95/Dlg/ZO-1) containing proteins through its PDZ binding motif (PBM). PTZ domains have crucial role in formation and function of signal transduction. Hippocampus is one of the cerebral regions that contain high load of viral antigens. We examined impact of RVG expression in the dorsal hippocampus on aversive as well as spatial learning and memory performance in rats. Two microliter of the lentiviral vector (~108 T.U./ml) encoding RVG or ∆RVG (deleted PBM) genomes was microinjected into the hippocampal CA1. After 1 week, rat's brain was cross-sectioned and RVG/∆RVG-expressing neuronal cells were confirmed by fluorescent microscopy. Passive avoidance and spatial learning and memory were assessed in rats by Shuttle box and Morris water maze (MWM). In the shuttle box, both RVG and ∆RVG decreased the time spent in the dark compartment compared to control (p < 0.05). In MWM, RVG and ∆RVG did not affect the acquisition of spatial task. In the probe test, RVG-expressing rats spent more time in the target quadrant, and also reached the platform position sooner than control group (p < 0.05). Rats expressing ∆RVG significantly swam farther from the hidden platform than RVG group (p < 0.05). Our data indicate RVG expression in the hippocampus strengthens aversive and spatial learning and memory performance. The boosting effect on spatial but not avoidance memory is mediated through PBM.

Dysregulation of ribosome-related genes in ankylosing spondylitis: a systems biology approach and experimental method.

BACKGROUND: Ankylosing spondylitis (AS) is an autoimmune rheumatic disease. Few candidate gene associations have been reported for AS and the current understanding of its pathogenesis remains still poor. Thus, the exact mechanism of AS is needed to urgently be disclosed. The purpose of this study was to identify candidate genes involving in AS disease. METHODS AND RESULTS: GSE25101 publicly available microarray and GSE117769 RNA-seq datasets of AS patients were obtained for bioinformatics analyses. Gene set enrichment analysis showed that in the microarray dataset, the ribosome pathway was significantly up-regulated in AS compared with controls. Furthermore, some ribosomal components demonstrated overexpression in patients in the RNA-seq dataset. To confirm the findings, 20 AS patients and 20 matching controls were selected from the Rheumatology Research Center clinic, Shariati Hospital. PBMCs were separated from whole blood and RNA contents were extracted. Following the results of datasets analysis, the expression level of rRNA5.8S pseudogene, rRNA18S pseudogene, RPL23, RPL7, and RPL17 genes were measured through real-time PCR. Our findings showed dysregulation of rRNA5.8S and rRNA18S pseudogenes, and also the RPL17 gene in patients. CONCLUSION: Considering that genes involved in ribosome biogenesis contributed to some AS-associated biological processes as well as diseases that have comorbidities with AS, our results might advance our understanding of the pathological mechanisms of ankylosing spondylitis.

Transduction efficacy and retrograde movement of a lentiviral vector pseudotyped by modified rabies glycoprotein throughout the trisynaptic circuit of the rat hippocampus.

BACKGROUND: The trisynaptic circuit (entorhinal cortex-dentate gyrus-CA3-CA1) is a key unidirectional network in the hippocampus. Damage to the hippocampus interrupts this circuit and causes neurological disorders. Efficient delivery of therapeutic genes into this network is of great interest with respect to treating trisynaptic circuit pathologies. METHODS: We generated a lentivector system pseudotyped by a variant of rabies glycoprotein, FUG-B2. The efficiency of the vector in the retrograde transduction of the rat hippocampal neurons (i.e. the entorhinal cortex from the dentate gyrus, the dentate gyrus from CA3, and CA3 from CA1) was examined by direct injection of the vector into the dentate gyrus, CA3 and CA1. To distinguish transduction of the neuronal and glial cells, as well as selective retrograde gene transfer, double-staining of the green fluorescent protein (GFP) expressing cells with the specific neuron biomarker NeuN (neuronal nuclear protein) and the specific glia biomarker GFAP (glial fibrillary acidic protein) was performed across the network. RESULTS: The transgene was successfully introduced into the circuit. More than 80% of the neuronal and glial cells at the injection sites preserved GFP expression during the 2-month period after vector injection. Importantly, GFP was expressed selectively in almost 80.0% of the presynaptic neuronal cells by retrograde axonal transport of the vector. CONCLUSIONS: The FUG-B2-based vector system can efficiently introduce the transgene into the rat hippocampal neurons both directly and indirectly through retrograde monosynaptic movement. This efficient and long-lasting gene delivery might provide a tool for treating neurological disorders originating in hippocampal circuits.

Monophosphoryl Lipid A and Pam3Cys Prevent the Increase in Seizure Susceptibility and Epileptogenesis in Rats Undergoing Traumatic Brain Injury.

Five percent of all epilepsy cases are attributed to traumatic brain injury (TBI), which are known as post-traumatic epilepsy (PTE). Finding preventive strategies for PTE is valuable. Remarkable feature of TBI is activation of microglia and subsequent neuroinflammation, which provokes epileptogenesis. The toll-like receptor agonists monophosphoryl lipid A (MPL) and tri-palmitoyl-S-glyceryl-cysteine (Pam3Cys) are safe, well-tolerated and effective adjuvants existing in prophylactic human vaccines. We examined the impact of early injection of MPL and Pam3Cys to rats, on the rate of kindled seizures acquisition following TBI. Rats received a single dose (1 µg/rat) of MPL or Pam3Cys through intracerebroventricular injection. 5 days later, trauma was exerted to temporo-parietal cortex of rats by controlled cortical impact device. After 24 h, traumatic rats underwent amygdala kindling. Brain level of the inflammatory cytokine tumor necrosis factor-alpha (TNF-α) was also measured in traumatic rats by immunoblotting. Compared to non-traumatic (sham-operated) rats, traumatic rats showed three times lower seizure threshold (133 ± 5 µA vs. 416.3 ± 16 µA, p < 0.001); about three times less number of stimuli to become kindled (5 ± 1 vs. 14 ± 2, p < 0.01); longer duration of kindled seizure parameters including entire seizure behavior, generalized seizures, and afterdischarges (p < 0.001); and a two times increase in the TNF-α level. MPL and Pam3Cys did not change kindling rate and the seizure parameters in sham-operated rats. The MPL- and Pam3Cys-pretreated traumatic rats displayed seizure threshold, speed of kindling, and duration of kindled seizure parameters, similar to the non-traumatic rats. Pretreatment by MPL and Pam3Cys prevented the increase in TNF-α level by trauma. Given that MPL and Pam3Cys currently have clinical use as well-tolerated vaccines with reliable safety, they have the potential to be used in prevention of PTE.

Toll-Like Receptor 1/2 Postconditioning by the Ligand Pam3cys Tempers Posttraumatic Hyperexcitability, Neuroinflammation, and Microglial Response: A Potential Candidate for Posttraumatic Epilepsy.

Toll-like receptors (TLRs) are activated by endogenous molecules released from damaged cells and contribute to neuroinflammation following traumatic brain injury (TBI) and epilepsy. TLR1/2 agonist tri-palmitoyl-S-glyceryl-cysteine (Pam3cys) is a vaccine adjuvant with confirmed safety in humans. We assessed impact of TLR1/2 postconditioning by Pam3cys on epileptogenesis and neuroinflammation in male rats, 6, 24, and 48 h after mild-to-moderate TBI. Pam3cys was injected into cerebral ventricles 30 min after controlled cortical impact (CCI) injury. After 24 h, rats underwent chemical kindling by once every other day injections of pentylenetetrazole (PTZ) 35 mg/kg until development of generalized seizures. Number of intact neurons, brain expression of proinflammatory cytokine TNF-α, anti-inflammatory cytokine IL-10, and marker of anti-inflammatory microglia arginase1 (Arg1) were determined by immunoblotting. Astrocytes and macrophage/microglia activation/polarization at the contused area was assessed by double immunostaining with Iba1/Arg1, Iba1/iNOS and GFAP/iNOS, specific antibodies. The CCI-injured rats became kindled by less number of PTZ injections than sham-operated rats (9 versus 14 injections, p < 0.0001). Pam3cys treatment returned the accelerated rate of epileptogenesis in TBI state to the sham level. Pam3cys decreased neural death 48 h after TBI. It decreased TNF-α (6 h post-TBI, p < 0.01), and up-regulated IL-10 (p < 0.01) and Arg1 (p < 0.05) 48 h after TBI. The iNOS-positive cells decreased (p < 0.001) whereas Iba1/Arg1-positive cells enhanced (p < 0.01) after Pam3cys treatment. Pam3cys inhibits TBI-accelerated acquisition of seizures. Pam3cys reprograms microglia and up-regulates anti-inflammatory cytokines during the first few days after TBI. This capacity along with the clinical safety, makes Pam3cys a potential candidate for development of effective medications against posttraumatic epilepsy.

Mitochondrial plasticity and synaptic plasticity crosstalk; in health and Alzheimer's disease.

Synaptic plasticity is believed to underlie the cellular and molecular basis of memory formation. Mitochondria are one of the main organelles involved in metabolism and energy maintenance as plastic organelles that change morphologically and functionally in response to cellular needs and regulate synaptic function and plasticity through multiple mechanisms, including ATP generation, calcium homeostasis, and biogenesis. An increased neuronal activity enhances synaptic efficiency, during which mitochondria's spatial distribution and morphology change significantly. These organelles build up in the pre-and postsynaptic zones to produce ATP, which is necessary for several synaptic processes like neurotransmitter release and recycling. Mitochondria also regulate calcium homeostasis by buffering intracellular calcium, which ensures proper synaptic activity. Furthermore, mitochondria in the presynaptic terminal have distinct morphological properties compared to dendritic or postsynaptic mitochondria. This specialization enables precise control of synaptic activity and plasticity. Mitochondrial dysfunction has been linked to synaptic failure in many neurodegenerative disorders, like Alzheimer's disease (AD). In AD, malfunctioning mitochondria cause delays in synaptic vesicle release and recycling, ionic gradient imbalances, and mostly synaptic failure. This review emphasizes mitochondrial plasticity's contribution to synaptic function. It also explores the profound effect of mitochondrial malfunction on neurodegenerative disorders, focusing on AD, and provides an overview of how they sustain cellular health under normal conditions and how their malfunction contributes to neurodegenerative diseases, highlighting their potential as a therapeutic target for such conditions.

Interleukin 4 Reduces Brain Hyperexcitability after Traumatic Injury by Downregulating TNF-α, Upregulating IL-10/TGF-ß, and Potential Directing Macrophage/Microglia to the M2 Anti-inflammatory Phenotype.

Macrophage/microglia are activated after Traumatic brain injury (TBI), transform to inflammatory phenotype (M1) and trigger neuroinflammation, which provokes epileptogenesis. Interleukin-4 (IL-4) is a well-known drive of macrophage/microglia to the anti-inflammatory phenotype (M2). We tested effect of IL-4 on speed of epileptogenesis, brain expression of inflammatory and anti-inflammatory cytokines, and lesion size in TBI-injured male rats. Rats underwent TBI by Controlled Cortical Impact. Then 100 ng IL-4 was injected into cerebral ventricles. One day after TBI, pentylenetetrazole (PTZ) kindling started and development of generalized seizures was recorded. The lesion size, cell survival rate, TNF-α, TGF-ß, IL-10, and Arginase1 (Arg1) was measured in the brain 6 h, 12 h, 24 h, 48 h, and 5 days after TBI. Astrocytes and macrophage/microglia activation/polarization was assessed by GFAP/Arg1 and Iba1/Arg1 immunostaining. TBI-injured rats were kindled by 50% less PTZ injections than control and sham-operated rats. IL-4 did not change kindling rate in sham-operated rats but inhibited acceleration of kindling rate in the TBI-injured rats. IL-4 decreased damage volume and number of destroyed neurons. IL-4 stopped TNF-α whereas upregulated TGF-ß, IL-10, and Arg1 expressions. Iba1/Arg1 positive macrophage/microglia was notably increased 48 h after IL-4 administration. IL-4 suppresses TBI-induced acceleration of epileptogenesis in rats by directing TBI neuroinflammation toward an anti-inflammatory tone and inhibition of cell death.

Latent toxoplasmosis impairs learning and memory yet strengthens short-term and long-term hippocampal synaptic plasticity at perforant pathway-dentate gyrus, and Schaffer collatterals-CA1 synapses.

Investigating long-term potentiation (LTP) in disease models provides essential mechanistic insight into synaptic dysfunction and relevant behavioral changes in many neuropsychiatric and neurological diseases. Toxoplasma (T) gondii is an intracellular parasite causing bizarre changes in host's mind including losing inherent fear of life-threatening situations. We examined hippocampal-dependent behavior as well as in vivo short- and long-term synaptic plasticity (STP and LTP) in rats with latent toxoplasmosis. Rats were infected by T. gondii cysts. Existence of REP-529 genomic sequence of the parasite in the brain was detected by RT-qPCR. Four and eight weeks after infection, spatial, and inhibitory memories of rats were assessed by Morris water maze and shuttle box tests, respectively. Eight weeks after infection, STP was assessed in dentate gyrus (DG) and CA1 by double pulse stimulation of perforant pathway and Shaffer collaterals, respectively. High frequency stimulation (HFS) was applied to induce LTP in entorhinal cortex-DG (400 Hz), and CA3-CA1 (200 Hz) synapses. T. gondii infection retarded spatial learning and memory performance at eight weeks post-infection period, whereas inhibitory memory was not changed. Unlike uninfected rats that normally showed paired-pulse depression, the infected rats developed paired-pulse facilitation, indicating an inhibitory synaptic network disruption. T. gondii-infected rats displayed strengthened LTP of both CA1-pyramidal and DG-granule cell population spikes. These data indicate that T. gondii disrupts inhibition/excitation balance and causes bizarre changes to the post-synaptic neuronal excitability, which may ultimately contribute to the abnormal behavior of the infected host.

Role of amyloid beta (25-35) neurotoxicity in the ferroptosis and necroptosis as modalities of regulated cell death in Alzheimer's disease.

Neuronal cell death as a prominent pathological feature contributes to cognitive decline and memory loss in Alzheimer's disease. We investigated the role of two forms of cell death pathways, ferroptosis and necroptosis, and their interactions following entorhinal cortex (EC) amyloidopathy. The Aß25-35 was bilaterally injected into the rat's EC, and Morris Water Maze was applied to determine spatial performance one week after Aß injection. For evaluation of ferroptosis and necroptosis involvement in Aß induced pathology, ferroptosis inhibitor, Ferrostatin (Fer-1), and necroptosis inhibitor, Necrostatin (Nec-1), were injected into the EC during training days of behavioral test. Our behavioral and histological assessment showed spatial learning and memory impairment, along with neuropathology changes such as cell survival and intracellular Aß deposits in response to EC amyloidopathy, which were ameliorated by treatment with Fer-1 or Nec-1. The expression of ferroptosis key factors GPX4 and SLC7A11 were decreased and the level of TfR was increased following Aß toxicity. Also, Necroptosis pathway related factors RIP1, RIP3, and MLKL were modulated by Aß neurotoxicity. However, application of Fer-1 or Nec-1 could inhibit the hippocampal ferroptosis and necroptosis pathways due to EC amyloidopathy. Our data also demonstrated that Aß-induced necroptosis suppressed by Fer-1, although Nec-1 had no effect on ferroptosis, indicating that ferroptosis pathway is upstream of necroptosis process in the Aß neurotoxicity. Moreover, Aß induced hippocampal mGLUR5 overexpression and reduced level of STIM1/2 recovered by Fer-1 or Nec-1. According to our findings ferroptosis and necroptosis pathways are involved in Aß neurotoxicity through modulation of mGLUR5 and STIM1/2 signaling.

Activating toll-like receptor 4 after traumatic brain injury inhibits neuroinflammation and the accelerated development of seizures in rats.

Toll-like receptor 4 (TLR4) signaling plays a detrimental role in traumatic brain injury (TBI) pathology. Pharmacologic or genetic inactivating TLR4 diminish TBI inflammation and neurological complications. Nonetheless, TLR4 priming alleviates TBI inflammation and seizure susceptibility. We investigated impact of postconditioning with TLR4 agonist monophosphoryl lipid A (MPL) on TBI neuroinflammation and epileptogenesis in rats. TBI was induced in temporo-parietal cortex of rats by Controlled Cortical Impact device. Then rats received a single dose (0.1 µg/rat) of MPL by intracerebroventricular injection. After 24 h, CCI-injured rats received intraperitoneal injection of pentylenetetrazole 35 mg/kg once every other day until acquisition of generalized seizures. The injury size, number of survived neurons, and brain protein level of TNF-α, TGF-ß, IL-10, and arginase1 (Arg1) were determined. Astrocytes and macrophage/microglia activation/polarization was assessed by double immunostaining with anti GFAP/Arg1 or anti Iba1/Arg1 antibodies. The CCI-injured rats developed generalized seizures after 5.9 ± 1.3 pentylenetetrazole injections (p < 0.001, compared to 12.3 ± 1.4 injections for sham-operated rats). MPL treatment returned the accelerated rate of epileptogenesis in TBI state to the sham-operated level. MPL did not change damage volume but attenuated number of dead neurons (p < 0.01). MPL decreased TNF-α overexpression (6 h post-TBI p < 0.0001), upregulated expression of TGF-ß (48 h post-TBI, p < 0.0001), and IL-10 (48 h post-TBI, p < 0.0001) but did not change Arg1 expression. GFAP/Arg1 and Iba1/Arg1 positive cells were detected in TBI area with no significant change following MPL administration. MPL administration after TBI reduces vulnerability to seizure acquisition through down regulating neural death and inflammation, and up-regulating anti-inflammatory cytokines. This capacity along with the clinical safety, makes MPL a potential candidate for development of drugs against neurological deficits of TBI.

Role of the glycoprotein thorns in anxious effects of rabies virus: Evidence from an animal study.

Rabies is a lethal infectious disease caused by rabies virus (RABV). Fear and anxiety are the distinguished symptoms in rabies patients. Fusion of RABV envelope glycoprotein (RVG) to host cell membrane initiates rabies pathogenesis via interacting with PDZ domain of signaling proteins. We assessed the anxiety-like behaviors, and hypothalamic-pituitary-adrenal axis (HPA) response to RVG infection. Contribution of PDZ binding motif (PBM) of RVG to the observed effects was also examined using a mutant form of RVG, ΔRVG, with deleted last four amino acids at PBM C-terminus. Lentiviral vectors containing RVG and/or ΔRVG genes were injected into the rat brain areas involved in anxiety including hypothalamus, dorsal hippocampus, and amygdala. RVG/ΔRVG neural expression was examined by fluorescent microscopy. Anxiety-like behaviors were assessed by elevated plus maze (EPM) and open field (OF) tasks. HPA response was evaluated via measuring corticosterone serum level by ELISA technique. RVG/ΔRVG were successfully expressed in neurons of the injected areas. RVG, but not ΔRVG, infection of hypothalamus and amygdala increased the time spent in EPM open arms, and OF total distance moved and velocity. RVG, but not ΔRVG, infection of hypothalamus and dorsal hippocampus increased corticosterone level. The anxiety-like behaviors and exploratory/locomotor activities of rats with RVG infection in hypothalamus, and amygdala are mediated by PBM of RVG. The HPA response to RVG infection of hypothalamus and dorsal hippocampus is dependent to PBM of RVG. Triggering anxiety-related signaling by PBM of RVG seems to be one of the mechanisms involved in anxiety behaviors seen in patients with rabies.

Phosphonate analog of 2-oxoglutarate regulates glutamate-glutamine homeostasis and counteracts amyloid beta induced learning and memory deficits in rats.

BACKGROUND: Metabolic alteration is a mainstream concept underlying the cognitive decline in neurodegenerative disorders including Alzheimer's disease (AD). Mitochondrial enzyme α-ketoglutarate dehydrogenase complex (α-KGDHC) seems to play a dual-edged sword role in cytotoxic insult. Here, using succinyl phosphonate (SP), a specific α-KGDHC inhibitor, we aimed to examine its potential action on AD progression. METHODS: Male Wistar rats were assigned to two separate experiments. First, they were bilaterally microinjected into the dorsal CA1 area by amyloid-beta (Aß)25-35 for four consecutive days. Seven days after the last injection, they were trained to acquire Morris Water Maze (MWM) task for three successive days when they were treated with SP after each training session. In the second experiment, SP was administered 30 min after the first Aß microinjection and behavioral tests were performed one week after the last Aß administration. The activity of glutamate dehydrogenase (GDH), and glutamine synthetase (GS), as key enzymes involved in glutamate-glutamine homeostasis and histological assays were evaluated in the hippocampi. RESULTS: Our behavioral results indicated that post-training SP treatment enhanced task acquisition but did not change memory performance in Aß-treated rats. However, administration of SP at the time of Aß injection precludes the deteriorative effect of Aß and neuronal injury on both spatial learning and memory performances indicating its preventive action against Aß pathology at its early stages. Measurement of enzymes activity shows that α-KGDHC activity was reduced in the Aß treated group, and SP administration restored its activity; also, GDH and GS activities were increased and decreased respectively due to Aß, and SP reversed the action of Aß on these enzymes. CONCLUSIONS: This study proposes that SP possibly a promising therapeutic approach to improve memory impairment in AD, especially in the early phases of this disease.

Anti-mullerian hormone attenuates insulin resistance and systemic inflammation in old obese C57BL/6 male mice.

PURPOSE: Epidemiological studies show that Anti-mullerian hormone (AMH) is inversely correlated with age, obesity-related diseases, and all-cause mortality in men. To further investigate the role of AMH in aging and obesity, we studied the effect of AMH treatment on the inflammatory and metabolic parameters and weight in old male C57BL/6 mice. METHOD: Thirty-six old male C57BL/6 mice (18 month-old) were either on the High-Fat Diet (HFD) or Normal Diet (ND). When obesity occurred in the HFD group, each group was divided into two subgroups; AMH-treated (ND+AMH and HFD+AMH) or controls (ND and HFD). The AMH subgroup received 15 ng/gbw of recombinant AMH injection every 48 h in four weeks. Then, serum AMH, CRP, fasting glucose, fasting insulin, and HOMA-IR were measured and analyzed. RESULTS: AMH injection decreased CRP level (HFD =622.86±25.73, HFD+AMH =543.2±24.99 ng/ml, p= 0.003), fasting insulin (HFD=1.50± 0.34, HFD+AMH =0.8±0.25 ng/ml, p=0.006) and HOMA-IR (HFD=12.76± 2.88, HFD+AMH =7.06±2.31, p=0.008) in the obese old mice comparison with control. In ND group, just CRP levels dropped following AMH injection (ND=451.24±20.61, ND+AMH= 326.8±23.76 ng/ml; p=0.001). Accelerated weight gain was observed in HFD+AMH compared with the HFD subgroup (p<0.05). CONCLUSIONS: In conclusion, increasing the circulating level of AMH could subside the systemic inflammation through decreasing CRP levels regardless of diet type and enhance insulin sensitivity in old obese mice. It can also lead to higher weight gain, without inflammation, in old obese male mice who are on an HFD.

Overexpression of protein kinase Mζ in the hippocampal dentate gyrus rescues amyloid-ß-induced synaptic dysfunction within entorhinal-hippocampal circuit.

Entorhinal cortex (EC) is one of the first cerebral regions affected in the early phase of Alzheimer's disease (AD). Soluble forms of amyloid beta (Aß) impair synaptic transmission in experimental AD models. Protein kinase Mζ (PKMζ) is an atypical persistently active protein kinase C, known to maintain long term synaptic plasticity and memory, but its role in AD has not yet been described. We examined effect of PKMζ overexpression on the late long-term potentiation (L-LTP) in the dentate gyrus (DG) following EC amyloidopathy. Oligomeric Aß 1-42 (oAß) or vehicle was bilaterally microinjected into the EC of the male Wistar rats. After 1 week, 2 µL of lentiviral vector (~108 TU/mL) encoding PKMζ genome was injected into the DG. One week later, synaptic responses and the LTP persistence were assessed in DG of freely moving animals during 90 minutes to 7 days period. Novel object recognition, passive avoidance and spatial memories were also tested. In rats with EC amyloidopathy, LTP was induced with less amplitude compared to the control group, and extinguished after 24 h. PKMζ overexpression in DG augmented synaptic responses (PS-LTP amplitudes) and maintained LTP over 1 week. PKMζ ameliorated recognition and memory deficits in rats with EC amyloidopathy. Microinjection of PKMζ inhibitor, zeta inhibitory peptide, into the DG abolished the boosting effect of PKMζ on synaptic activity and memory performance. PKMζ-dependent pathway could be a potential therapeutic target to combat synaptic failure and memory deficit in the early phase of AD.

Overexpression of protein kinase Mζ in the hippocampus mitigates Alzheimer's disease-related cognitive deficit in rats.

Accumulation of amyloid beta (Aß) soluble forms in the cerebral parenchyma is the mainstream concept underlying memory deficit in the early phase of Alzheimer's disease (AD). PKMζ plays a critical role in the maintenance of long-term memory. Yet, the role of this brain-specific enzyme has not been addressed in AD. We examined the impact of hippocampal PKMζ overexpression on AD-related memory impairment in rats. Oligomeric form of Aß (oAß) or vehicle was bilaterally microinjected into the dorsal hippocampus of male Wistar rats under stereotaxic surgery. One week later, 2 µl of lentiviral vector (108 T.U. / ml.) encoding PKMζ genome was microinjected into the dorsal hippocampus. Seven days later, behavioral performance was assessed using shuttle box and Morris water maze. The expression levels of GluA1, GluA2 and KCC2 were determined in the hippocampus using western blot technique. Our data showed that oAß impairs both passive avoidance and spatial learning and memory. However, overexpression of PKMζ in the dorsal hippocampus restored the behavioral performance. This improving effect was blocked by microinjection of ZIP, a PKMζ inhibitor, into the hippocampus. oAß or PKMζ did not significantly change GluA1 level in the hippocampus. Furthermore, PKMζ failed to restore elevated KCC2 level induced by oAß. However, oAß decreased GluA2 level, and overexpression of PKMζ restored its expression toward the control level. In conclusion, hippocampal overexpression of PKMζ restored memory dysfunction induced by amyloidopathy in part, through preserving hippocampal GluA2 containing AMPA receptors. PKMζ's signaling pathway could be considered as a therapeutic target to battle memory deficits in the early phase of AD.

Fabricating an electroactive injectable hydrogel based on pluronic-chitosan/aniline-pentamer containing angiogenic factor for functional repair of the hippocampus ischemia rat model.

The hippocampus, a critical cerebral region involved in learning and memory formation, is especially vulnerable to ischemic defect. Here, we developed an injectable electroactive hydrogel based on pluronic-chitosan/aniline-pentamer with proper conductivity around 10-4 S/cm to achieve the functional repair of the hippocampus following the ischemic defect. FTIR, DSC, and TGA measurements were performed to assess the chemical structure and thermal stability of the synthesized hydrogel. Aniline pentamer decreased the swelling capacity, degradation, and drug release rate. Further, contact angle, melting point, and gelation time of hydrogels were enhanced by addition of aniline oligomer. Moreover, it endowed the on-demand electro-responsive drug release. Injectability of hydrogel was evaluated by rheometry, exhibiting proper gelling time at the body temperature. The ionic/electrical conductivity and desired in vitro biocompatibility with PC12 cells were also achieved. Injection of VEGF-loaded electroactive hydrogel in the hippocampal ischemic animal model resulted in decreased infarction volume, improved hippocampal dependent learning, and memory performance. Taken all together, the results confirmed that fabricated injectable hydrogel would be a suitable candidate for ischemic defect treatment and can lead to new horizons to treat neurological disorders.

Preconditioning with toll-like receptor agonists attenuates seizure activity and neuronal hyperexcitability in the pilocarpine rat model of epilepsy.

Neuroinflammation plays an important role in epileptic disorders. Toll-like receptors (TLRs) are the key signal transduction tools by which neuroinflammation may promote epileptogenesis. Depending on the stimulus nature, TLRs may engage a distinct signaling pathway. We examined the impact of early minor activation of TLR4 and TLR2 on the severity of seizure in the pilocarpine rat model of temporal lobe epilepsy (TLE). One µg of Lipopolysaccharides (LPS), Monophosphoryl lipid A (MPL), Pam3Cysor or vehicles were microinjected into the right lateral ventricle of the male Wistar rats. 24 h later, seizures were induced by intraperitoneal injection of pilocarpine, and seizure-related behaviors were monitored. 24 h after seizure induction, the hippocampal level of pro/anti-inflammatory mediators and electrophysiological properties of the dentate gyrus (DG) granular cells were investigated by western blot and whole cell patch clamp techniques, respectively. Pretreatment with TLR ligands resulted in decreased seizure severity, lower hippocampal pro-inflammatory (IL-1ß and IL-6) cytokines and higher anti-inflammatory (IL-10 and TGF- ß) mediators in the pilocarpine-treated rats. Pilocarpine induced profound hyperexcitability in the DG granule cells accompanied by potentiated excitatory postsynaptic currents (EPSCs) and dampened inhibitory postsynaptic currents (IPSCs), in contrast to the control group. However, pretreatment with TLR ligands preserved almost normal excitability and synaptic transmission against the pilocarpine. In conclusion, early activation of TLR4 and TLR2, probably through preserving normal hippocampal cytokine profile and neuronal function attenuates seizure severity in the rat model of TLE.