Pilocarpine

Peripheral myeloid cell EP2 activation contributes to the deleterious consequences of status epilepticus.[Pubmed:33293358]

J Neurosci. 2020 Dec 8. pii: JNEUROSCI.2040-20.2020.

A multidimensional inflammatory response ensues after status epilepticus (SE), driven partly by cyclooxygenase-2 mediated activation of prostaglandin EP2 receptors. The inflammatory response is typified by astrocytosis, microgliosis, erosion of the blood-brain barrier (BBB), formation of inflammatory cytokines, and brain infiltration of blood-borne monocytes. Our previous studies have shown that inhibition of monocyte brain invasion or systemic administration of an EP2 receptor antagonist relieves multiple deleterious consequences of SE. Here we identify those effects of EP2 antagonism that are reproduced by conditional ablation of EP2 receptors in immune myeloid cells and show that systemic EP2 antagonism blocks monocyte brain entry in male mice. The induction of hippocampal IL-6 after Pilocarpine SE was nearly abolished in EP2 conditional knockout mice. Serum albumin levels in the cortex, a measure of BBB breakdown, were significantly higher after SE in EP2-sufficient mice but not in EP2 conditional knockouts. EP2 deficiency in innate immune cells accelerated the recovery from sickness behaviors following SE. Surprisingly, neurodegeneration was not alleviated in myeloid conditional knockouts. Systemic EP2 antagonism prevented monocyte brain infiltration and provided broader rescue of SE-induced effects than myeloid EP2 ablation, including neuroprotection and broader suppression of inflammatory mediators. Reporter expression indicated the cellular target of CD11b-driven Cre was circulating myeloid cells but, unexpectedly, not microglia. These findings indicate that activation of EP2 receptors on immune myeloid cells drives substantial deficits in behavior and disrupts the BBB after SE. The benefits of systemic EP2 antagonism can be attributed, in part, to blocking brain recruitment of blood-borne monocytes.SIGNIFICANCE STATEMENTUnabated seizures reduce quality of life, promote the development of epilepsy, and can be fatal. We previously identified activation of prostaglandin EP2 receptors as a driver of undesirable consequences of seizures. However, the relevant EP2-expressing cell types remain unclear. Here we identify peripheral innate immune cells as a driver of the EP2-related negative consequences of seizures. Removal of EP2 from peripheral immune cells was beneficial, abolishing production of a key inflammatory cytokine, accelerating weight regain, and limiting behavioral deficits. These findings provide evidence that EP2 engagement on peripheral immune and brain endothelia contributes to the deleterious effects of SE, and will assist in the development of beneficial therapies to enhance quality of life in individuals who suffer prolonged seizures.

Lrp4 in hippocampal astrocytes serves as a negative feedback factor in seizures.[Pubmed:33292473]

Cell Biosci. 2020 Nov 23;10(1):135.

BACKGROUND: Epilepsy is characterized by the typical symptom of seizure, and anti-seizure medications are the main therapeutic method in clinical, but the effects of these therapy have not been satisfactory. To find a better treatment, it makes sense to further explore the regulatory mechanisms of seizures at genetic level. Lrp4 regionally expresses in mice hippocampus where is key to limbic epileptogenesis. It is well known that neurons release a high level of glutamate during seizures, and it has been reported that Lrp4 in astrocytes down-regulates glutamate released from neurons. However, it is still unclear whether there is a relationship between Lrp4 expression level and seizures, and whether Lrp4 plays a role in seizures. RESULTS: We found that seizures induced by Pilocarpine decreased Lrp4 expression level and increased miR-351-5p expression level in mice hippocampus. Glutamate reduced Lrp4 expression and enhanced miR-351-5p expression in cultured hippocampal astrocytes, and these effects can be partially attenuated by AP5. Furthermore, miR-351-5p inhibitor lessened the reduction of Lrp4 expression in glutamate treated hippocampal astrocytes. Local reduction of Lrp4 in hippocampus by sh Lrp4 lentivirus injection in hippocampus increased the threshold of seizures in Pilocarpine or pentylenetetrazol (PTZ) injected mice. CONCLUSIONS: These results indicated that high released glutamate induced by seizures down-regulated astrocytic Lrp4 through increasing miR-351-5p in hippocampal astrocytes via activating astrocytic NMDA receptor, and locally reduction of Lrp4 in hippocampus increased the threshold of seizures. Lrp4 in hippocampal astrocytes appears to serve as a negative feedback factor in seizures. This provides a new potential therapeutic target for seizures regulation.

Status epilepticus induced Gadd45b is required for augmented dentate neurogenesis.[Pubmed:33279798]

Stem Cell Res. 2020 Nov 30;49:102102.

In animal models with temporal lobe epilepsy (TLE), the status epilepticus (SE) leads to a dramatic increase in number of newly born neuron in the subgranular zone (SGZ) of dentate gyrus. How the SE confers a modulation in the dentate neurogenesis is mostly unknown. Gadd45b is involved in epigenetic gene activation by DNA demethylation. This study was performed to present a novel mechanism underling SE-induced dentate neurogenesis. A transient induction (12 hrs to 3 days) of Gadd45b was observed in dentate gyrus of mice after Pilocarpine-induced SE. Labeling the dividing cells with BrdU, we next found that the induction of Gadd45b was required to increase the rate of cell proliferation in the dentate gyrus at 7 and 14 days after SE. Afterward, the DNA methylation levels for candidate growth factor genes critical for the adult neurogenesis were assayed with Sequenom MassARRAY Analyzer. The results indicated that Gadd45b was necessary for SE-induced DNA demethylation of specific promoters and expression of corresponding genes in the dentate gyrus, including brain-derived neurotrophic factor (BDNF) and fibroblast growth factor-2 (FGF-2). Using Timm staining, we further suggested that SE-induced Gadd45b might contribute to the subsequent mossy fiber sprouting (MFS) in the chronically epileptic hippocampus via epigenetic regulation of dentate neurogenesis at early stage after SE. Together, Gadd45b links Pilocarpine-induced SE to epigenetic DNA modification of secreted factors in the dentate gyrus, leading to extrinsic modulation on the neurogenesis.

Antifungal activity and antidiarrheal activity via antimotility mechanisms of (-)-fenchone in experimental models.[Pubmed:33268962]

World J Gastroenterol. 2020 Nov 21;26(43):6795-6809.

BACKGROUND: (-)-Fenchone is a bicyclic monoterpene present in essential oils of plant species, such as Foeniculum vulgare and Peumus boldus, used to treatment of gastrointestinal diseases. Pharmacological studies report its anti-inflammatory, antioxidant, and antinociceptive activity. AIM: To investigate antidiarrheal activity related to gastrointestinal motility, intestinal secretion and antimicrobial activity. METHODS: A castor oil-induced diarrhea model was used to evaluate antidiarrheal activity. Intestinal transit and gastric emptying protocols were used to assess a possible antimotility effect. Muscarinic receptors, presynaptic alpha2-adrenergic and tissue adrenergic receptors, KATP channels, nitric oxide were investigated to uncover antimotility mechanisms of action and castor oil-induced enteropooling to elucidate antisecretory mechanisms. The antimicrobial activity was evaluated in the minimum inhibitory concentration model, the fractional inhibitory concentration index using the (-)-fenchone association method with standard antifungal agents. RESULTS: (-)-Fenchone (75, 150 and 300 mg/kg) showed antidiarrheal activity, with a significant decrease in the evacuation index. This activity is possibly related to a percentage of reduced intestinal transit (75, 150 and 300 mg/kg). The antimotility effect of (-)-fenchone decreased in the presence of Pilocarpine, yohimbine, propranolol, L-NG-nitroarginine methyl ester or glibenclamide. In the enteropooling model, no reduction in intestinal fluid weight was observed. (-)- Fenchone did not show antibacterial activity; on the other hand, inhibits the growth of strains of fungi with a minimum fungicide concentration of 32 mug/mL. However, when it was associated with amphotericin B, no synergism was observed. CONCLUSION: The antidiarrheal effect of (-)-fenchone in this study involves antimotility effect and not involve antisecretory mechanisms. (-)-Fenchone presents antifungal activity; however, it did not show antibacterial activity.

Hormonal and biochemical changes in female Proechimys guyannensis, an animal model of resistance to pilocarpine-induced status epilepticus.[Pubmed:33268798]

Sci Rep. 2020 Dec 2;10(1):20982.

The Amazon rodent Proechimys guyannensis is widely studied for hosting various pathogens, though rarely getting sick. Previous studies on male Proechimys have revealed an endogenous resistance to epilepsy. Here, we assess in female Proechimys, whether sex hormones and biochemical aspects can interfere with the induction of status epilepticus (SE). The lithium-Pilocarpine ramp-up protocol was used to induce SE, and blood sera were collected at 30 and 90 min after SE, alongside brains, for biochemical, western blot and immunohistochemical analyses. Results from non-ovariectomised (NOVX) Proechimys were compared to ovariectomised (OVX) animals. Data from female Wistars were used as a positive control of SE inductions. SE latency was similar in NOVX, OVX, and female Wistars groups. However, the Pilocarpine dose required to induce SE in Proechimys was higher (25- to 50-folds more). Despite a higher dose, Proechimys did not show strong SE like Wistars; they only reached stage 2 of the Racine scale. These data suggest that female Proechimys are resistant to SE induction. Glucose and progesterone levels increased at 30 min and returned to normal at 90 min after SE. A relevant fact because in humans and rodents, SE leads to hypoglycaemia after 30 min of SE and does not return to normal levels in a short time, a typical adverse effect of SE. In OVX animals, a decrease in GABAergic receptors within 90 min of SE may suggest that ovariectomy produces changes in the hippocampus, including a certain vulnerability to seizures. We speculate that progesterone and glucose increases form part of the compensatory mechanisms that provide resistance in Proechimys against SE induction.

Long-lasting tagging of neurons activated by seizures or cocaine administration in Egr1-CreER(T2) transgenic mice.[Pubmed:33226686]

Eur J Neurosci. 2020 Nov 23.

Permanent tagging of neuronal ensembles activated in specific experimental situations is an important objective to study their properties and adaptations. In the context of learning and memory these neurons are referred to as engram neurons. Here we describe and characterize a novel mouse line, Egr1-CreER(T2) , which carries a transgene in which the promoter of the immediate early gene Egr1 drives the expression of the CreER(T2) recombinase that is only active in the presence of tamoxifen metabolite, 4-hydroxy-tamoxifen (4-OHT). Egr1-CreER(T2) mice were crossed with various reporter mice, Cre-dependently expressing a fluorescent protein. Without tamoxifen or 4-OHT, no or few tagged neurons were observed. Epileptic seizures induced by Pilocarpine or pentylenetetrazol in the presence of tamoxifen or 4-OHT, induced the persistent tagging of many neurons and some astrocytes in the dentate gyrus of hippocampus, where Egr1 is transiently induced by seizures. One week after cocaine and 4-OHT administration, these mice displayed a higher number of tagged neurons in the dorsal striatum than saline/4-OHT controls, with differences between reporter lines. Cocaine-induced tagging required ERK activation and tagged neurons were more likely than others to exhibit ERK phosphorylation or Fos induction after a second injection. Interestingly neurons tagged in saline-treated mice also had an increased propensity to express Fos, suggesting the existence of highly responsive striatal neurons susceptible to be re-activated by cocaine repeated administration, which may contribute to the behavioral adaptations. Our report validates a novel transgenic mouse model for permanently tagging activated neurons and studying long term alterations of Egr1-expressing cells.

Bioinformatic analysis identifies key transcriptome signatures in temporal lobe epilepsy.[Pubmed:33225612]

CNS Neurosci Ther. 2020 Dec;26(12):1266-1277.

AIMS: To identify transcriptome signatures underlying epileptogenesis in temporal lobe epilepsy (TLE). METHODS: Robust rank aggregation analysis was used to integrate multiple microarrays in rodent models of TLE and determine differentially expressed genes (DEGs) in acute, latent, and chronic stages. Functional annotation and protein-protein interaction analysis were performed to explore the potential functions of the DEGs and identify hub genes with the highest intramodular connectivity. The association between hub genes and hippocampal sclerosis/seizure frequency was analyzed using publicly available RNA-sequencing datasets from TLE patients. We subsequently established a Pilocarpine-induced status epilepticus (SE) model in rats and validated mRNA expression of hub genes by quantitative reverse transcription PCR (qRT-PCR). RESULTS: The DEGs in the acute, latent, and chronic phases of TLE in animal models were prominently enriched in inflammatory response. Hub genes identified in the acute phase mainly participated in biological processes including inflammation, blood-brain barrier damage, and cell adhesion. The hub genes in the latent phase were related to microglia/macrophage activation (Emr1 and Aif1) and phagocytosis (Cd68, Tyrobp, and Lyz). In the chronic phase, the hub genes were associated with activation of complements and microglia/macrophages. We further found that some hub genes identified in human TLE, such as Tlr2, Lgals3, and Stat3, were positively correlated with seizure frequency. Other hub genes, including Lgals3 and Serpine1, were associated with hippocampus sclerosis. qRT-PCR analysis confirmed that the mRNA levels of hub genes in rat hippocampus were significantly up-regulated after SE induction. CONCLUSIONS: Our integrated analysis identified hub genes in different stages of epilepsy. The functional annotations suggest that the activation and phagocytic activities of microglia/macrophages may play critical roles in epileptogenesis of TLE.

Hemopressin as a breakthrough for the cannabinoid field.[Pubmed:33212113]

Neuropharmacology. 2020 Nov 16;183:108406.

Hemopressin (PVNFKFLSH in rats, and PVNFKLLSH in humans and mice), a fragment derived from the alpha-chain of hemoglobin, was the first peptide described to have type 1 cannabinoid receptor activity. While hemopressin was shown to have inverse agonist/antagonistic activity, extended forms of hemopressin (i.e. RVD-hemopressin, also called pepcan-12) exhibit type 1 and type 2 cannabinoid receptor agonistic/allosteric activity, and recent studies suggest that they can activate intracellular mitochondrial cannabinoid receptors. Therefore, hemopressin and hemopressin-related peptides could have location-specific and biased pharmacological action, which would increase the possibilities for fine-tunning and broadening cannabinoid receptor signal transduction. Consistent with this, hemopressins were shown to play a role in a number of physiological processes including antinociceptive and anti-inflammatory activity, regulation of food intake, learning and memory. The shortest active hemopressin fragment, NFKF, delays the first seizure induced by Pilocarpine, and prevents neurodegeneration in an experimental model of autoimmune encephalomyelitis. These functions of hemopressins could be due to engagement of both cannabinoid and non-cannabinoid receptor systems. Self-assembled nanofibrils of hemopressin have pH-sensitive switchable surface-active properties, and show potential as inflammation and cancer targeted drug-delivery systems. Upon disruption of the self-assembled hemopressin nanofibril emulsion, the intrinsic analgesic and anti-inflammatory properties of hemopressin could help bolster the therapeutic effect of anti-inflammatory or anti-cancer formulations. In this article, we briefly review the molecular and behavioral pharmacological properties of hemopressins, and summarize studies on the intricate and unique mode of generation and binding of these peptides to cannabinoid receptors. Thus, the review provides a window into the current status of hemopressins in expanding the repertoire of signaling and activity by the endocannabinoid system, in addition to their new potential for pharmaceutic formulations.

Unilateral Toxic Anterior Segment Syndrome Resulting in Cataract and Urrets-Zavalia Syndrome after Sequential Uneventful Implantation of a Posterior Chamber Phakic Toric Intraocular Lens at Two Different Surgical Facilities: A Series of Unfortunate Events.[Pubmed:33204556]

Case Rep Ophthalmol Med. 2020 Nov 3;2020:1216578.

Background: Phakic Intraocular Lens (P-IOL) implantation is a safe, easy, predictable intervention designed to manage moderate to high refractive errors. Complications are relatively uncommon and include mainly cataract and intraocular pressure spikes. Toxic Anterior Segment Syndrome (TASS) is a rather unusual sterile anterior segment inflammation after uneventful intraocular surgery, extremely rarely reported after P-IOL implantation. Urrets-Zavalia Syndrome (UZS) is also very rarely described after P-IOL. To date, to the best of the authors' knowledge, no article has ever described the simultaneous occurrence of TASS and UZS in a patient after P-IOL implantation. Objective: In this article, the authors present the case of a female patient with moderate myopic astigmatism, who underwent sequential P-IOL implantation at two different facilities. The postoperative course of the first eye was uneventful, but she developed complications associated to the intervention in the second eye. Materials: The article describes the case of a young patient who underwent a sequential Phakic Intraocular Lens (P-IOL) implantation at two different institutions. The postoperative course of the first eye (left eye) was uneventful; however, the second eye (right eye) initially developed Toxic Anterior Segment Syndrome (TASS). Although timely and correct management was instituted, upon resolution of TASS, the patient developed Urrets-Zavalia Syndrome, anterior subcapsular cataract, and significant endothelial damage in the same eye. Results: The patient was followed closely and managed accordingly; corneal edema and anterior segment inflammation of the right eye eventually resolved. Nevertheless, an anterior subcapsular cataract and a fixed dilated pupil remained; with normal intraocular pressure (IOP). Specular microscopy confirmed an endothelial cell loss in the TASS eye (right eye). Pupil size showed no reaction to repeated doses of Pilocarpine 2%. A month after surgery, refraction on her right eye was +0.25 + 0.75 x 93, which resulted in a 20/50 vision. Conclusions: TASS and UZS are both extremely rare complications after uneventful P-IOL implantation, with only a handful of cases having been reported of each of them. To date, this is the very first case where UZS ensued after and potentially as a consequence of TASS in a patient who had undergone P-IOL implantation. Although a direct causative element could not be pinpointed, the fact that the complication ensued after being operated in one surgical institution and not the other, could suggest some role of different sterilization and handling procedures, but no direct conclusion can be made on this case.

Amygdala Low-Frequency Stimulation Reduces Pathological Phase-Amplitude Coupling in the Pilocarpine Model of Epilepsy.[Pubmed:33202818]

Brain Sci. 2020 Nov 13;10(11). pii: brainsci10110856.

Temporal-lobe epilepsy (TLE) is the most common type of drug-resistant epilepsy and warrants the development of new therapies, such as deep-brain stimulation (DBS). DBS was applied to different brain regions for patients with epilepsy; however, the mechanisms of action are not fully understood. Therefore, we tried to characterize the effect of amygdala DBS on hippocampal electrical activity in the lithium-Pilocarpine model in male Wistar rats. After status epilepticus (SE) induction, seizure patterns were determined based on continuous video recordings. Recording electrodes were inserted in the left and right hippocampus and a stimulating electrode in the left basolateral amygdala of both Pilo and age-matched control rats 10 weeks after SE. Daily stimulation protocol consisted of 4 x 50 s stimulation trains (4-Hz, regular interpulse interval) for 10 days. The hippocampal electroencephalogram was analyzed offline: interictal epileptiform discharge (IED) frequency, spectral analysis, and phase-amplitude coupling (PAC) between delta band and higher frequencies were measured. We found that the seizure rate and duration decreased (by 23% and 26.5%) and the decrease in seizure rate correlated negatively with the IED frequency. PAC was elevated in epileptic animals and DBS reduced the pathologically increased PAC and increased the average theta power (25.9% +/- 1.1 vs. 30.3% +/- 1.1; p < 0.01). Increasing theta power and reducing the PAC could be two possible mechanisms by which DBS may exhibit its antiepileptic effect in TLE; moreover, they could be used to monitor effectiveness of stimulation.

Rosiglitazone Prevents Autophagy by Regulating Nrf2-Antioxidant Response Element in a Rat Model of Lithium-pilocarpine-induced Status Epilepticus.[Pubmed:33197503]

Neuroscience. 2020 Nov 13. pii: S0306-4522(20)30687-4.

Status epilepticus (SE) leads to irreversible neuronal damage and consists of a complex pathogenesis that involves oxidative stress and subsequent autophagy. Rosiglitazone has recently been considered as a potential neuroprotective factor in epilepsy because of its antioxidative function. The aim of this study was to assess the effects of rosiglitazone in SE rat models and investigate whether its mechanisms of action involve autophagy via the antioxidant factor, nuclear factor erythroid 2-related factor 2 (Nrf2). The male Sprague-Dawley rats (200-220g) were used to establish lithium-Pilocarpine-induced SE model. We found that rosiglitazone markedly improved neuronal survival at 24-h post-SE as indicated via Hematoxylin-Eosin and Nissl staining. Furthermore, along with a reduction in reactive oxygen species, rosiglitazone pretreatment enhanced the antioxidative activity of superoxide dismutase and the expression level of Nrf2, as detected via chemical assay kits and Western blotting, respectively. In addition, the microtubule-associated protein light chain 3II (LC3II)/LC3I ratio was increased and peaked at 24h after SE, whereas p62 mRNA levels were sharply elevated at 72h after SE, both SE-induced increases of which were reversed via rosiglitazone pretreatment. To further test our hypothesis of the key role of Nrf2 in this process, small-interfering RNA for Nrf2 (siNrf2) was then transfected into SE rats to knockdown Nrf2 expression. We found that siNrf2 partially blocked the above effects of rosiglitazone on autophagy-related proteins in SE rats. Taken together, our findings suggest that rosiglitazone attenuates oxidative-stress-induced autophagy via increasing Nrf2 in SE rats and may be used as a promising therapeutic strategy for SE treatment.

d-Serine Intervention In The Medial Entorhinal Area Alters TLE-Related Pathology In CA1 Hippocampus Via The Temporoammonic Pathway.[Pubmed:33197499]

Neuroscience. 2020 Nov 14. pii: S0306-4522(20)30686-2.

Entrainment of the hippocampus by the medial entorhinal area (MEA) in Temporal Lobe Epilepsy (TLE), the most common type of drug-resistant epilepsy in adults, is believed to be mediated primarily through the perforant pathway (PP), which connects stellate cells in layer (L) II of the MEA with granule cells of the dentate gyrus (DG) to drive the hippocampal tri-synaptic circuit. Using immunohistochemistry, high-resolution confocal microscopy and the rat Pilocarpine model of TLE, we show here that the lesser known temporoammonic pathway (TAP) plays a significant role in transferring MEA pathology to the CA1 region of the hippocampus independently of the PP. The pathology observed was region-specific and restricted primarily to the CA1c subfield of the hippocampus. As shown previously, daily intracranial infusion of d-serine (100mum), an antagonist of GluN3-containing triheteromeric N-Methyl d-aspartate receptors (t-NMDARs), into the MEA prevented loss of LIII neurons and epileptogenesis. This intervention in the MEA led to the rescue of hippocampal CA1 neurons that would have otherwise perished in the epileptic animals, and down regulation of the expression of astrocytes and microglia thereby mitigating the effects of neuroinflammation. Interestingly, these changes were not observed to a similar extent in other regions of vulnerability like the hilus, DG or CA3, suggesting that the pathology manifest in CA1 is driven predominantly through the TAP. This work highlights TAP's role in the entrainment of the hippocampus and identifies specific areas for therapeutic intervention in dealing with TLE.

Isovaline efficacy in a rat pup model of infantile spasms.[Pubmed:33196548]

Neuroreport. 2021 Jan 6;32(1):61-65.

Infantile spasms, also known as epileptic spasms during infancy, is an epileptic disorder of infancy and early childhood that is associated with developmental delay or regression, high mortality rate and is difficult to treat with conventional antiseizure medication. Previously, we reported that a unique amino acid called isovaline had potent anticonvulsive efficacy in the 4-aminopyridine and Pilocarpine rat models of seizures. In this study, we examined whether isovaline possess therapeutic utility in a well-established rat model of infantile spasms which involves the pretreatment of a pregnant dam with betamethasone and subsequent induction of spasms with N-methyl-D-asparate (NMDA), a glutamate receptor agonist, in 15-day old pups. We treated seven of these pups with saline prior to administering NMDA and eight of these pups with isovaline (300 mg/kg) intraperitoneal (i.p.) prior to NMDA. Isovaline significantly reduced the number of full-body jumps from 18.1 +/- 5.0 to 6.3 +/- 1.8 and leg/arm/tail strains from 4.4 +/- 1.6 to 1.1 +/- 0.5. A trend in a reduction of body twitch was noted in rat pups administered isovaline (P = 0.05), but no significant difference was seen in NMDA-induced head nods (P = 0.221). In conclusion, our data demonstrate a potential for isovaline to attenuate an aggressive form of epilepsy that typically requires highly toxic medications to treat in children.

Proteomic Analysis of Exosome-Like Vesicles Isolated From Saliva of the Tick Haemaphysalis longicornis.[Pubmed:33194791]

Front Cell Infect Microbiol. 2020 Oct 22;10:542319.

Extracellular vesicles (EVs), are considered as vehicles of cellular communication. Parasites usually release EVs in their excretory-secretory products to modulate host environment. However, little is known about the secretion of EVs by ticks. In this study, we show for the first time that the tick Haemaphysalis longicornis secretes EVs in saliva that resembles exosomes. EVs were purified from Pilocarpine induced saliva of partially engorged H. longicornis ticks. Electron microscopy analysis revealed the presence of exosome-like vesicles with a size of 100 nm. Proteomic analysis by LC-MS/MS identified a total of 356 proteins in tick-derived EVs. Proteome data of tick-derived EVs was validated by Western blot analysis. Immunodetection of Hsp70 and GAPDH proteins indicated that the proteomics data of tick-derived EVs were highly reliable. Bioinformatics analysis (Gene Ontology) indicated association of certain biological and molecular functions with proteins which may be helpful during tick development. Likewise, KEGG database revealed involvement of vesicular proteins in proton transport, detoxification, ECM-receptor interaction, ribosome, RNA transport, ABC transporters, and oxidative phosphorylation. The results of this study provide evidence that EVs are being secreted in tick saliva and suggest that tick saliva-derived EVs could play important roles in host-parasite relationships. Moreover, EVs could be a useful tool in development of vaccines or therapeutics against ticks.

Time-Dependent Impairment of Fear Conditioning and Associated Brain Regions After Pilocarpine-Induced Status Epilepticus.[Pubmed:33192357]

Front Behav Neurosci. 2020 Aug 27;14:149.

Our study aimed to demonstrate time-dependent declarative memory changes and its associated brain regions after status epilepticus (SE) using structural imaging techniques and machine learning methods. Pilocarpine was administrated to establish the SE model. At four different time points after SE (1, 2, 3, and 4 months, respectively), rats were subjected to structural imaging acquisition as well as contextual fear conditioning for the measurement of brain structural changes and declarative memory. Voxel-based morphometry (VBM) analysis were performed. Those significantly different regions were selected as features for training support vector machine (SVM). A linear kernel was chosen for regression of declarative memory. Leave-one-out cross-validation was applied to ensure generalization. Our results showed that the Pilocarpine groups displayed the most severely impaired declarative memory at 2 months after SE and improved afterward, but failed to recover to the normal condition at 4 months after SE. The Pilocarpine groups showed lower gray matter volumes and larger cerebrospinal fluid (CSF) volumes. After controlling for the total brain volumes, ANOVA demonstrated gray matter volume changes in the CA1 subfield of the hippocampus, primary somatosensory cortex, entorhinal cortex, etc. The combination of VBM and SVM identifies the somatosensory cortex and entorhinal cortex as the correlated brain regions for declarative memory dysfunctions after SE. Our study indicates that compensational mechanisms might be triggered to help with the recovery of memory functions after SE. Structural changes of the somatosensory cortex and entorhinal cortex might be involved in memory impairment after SE.