Design and evaluation of pharmacological properties of a new 1,3,4-thiadiazolylamide derivative of 2-propylpentanoic acid

Introduction: The use of the pharmacophoric approach is a promising direction for modifying the chemical structure of 2-propylpentanoic (valproic) acid in order to obtain new drugs. Materials and methods: In the experiments on mice, acute toxicity, neurotoxicity, antiepileptic activity and analgesic effect of N-(5-ethyl-1,3,4-thiadiazol-2-yl)-2-propylpentanamide (valprazolamide) were evaluated. LD50 was determined by probit analysis. Neurotoxicity was determined in a rotarod test and a bar test in mice. The effects of valprazolamide on the exploratory behavior of mice in open field test and in a light/dark transition test were evaluated. Its antiepileptic activity was tested in mice against seizures induced by maximal electroshock, pentylenetetrazole (scPTZ); isoniazid, thiosemicarbazide, pilocarpine, and camphor. The analgesic effect was studied in a hot plate test. Results and discussion: N-(5-ethyl-1,3,4-thiadiazol-2-yl)-2-propylpentanamide was obtained by introducing pharmacophores into the structure of 2-propylpentanoic acid: a substituted amide group and an electron-donor domain of 1,3,4-thiadiazole. The LD50 value for intraperitoneal administration of a new 2-propylpentanoic acid: derivative to mice was 924.8 mg/kg, and the TD50 value in the rotarod test and the bar test were 456.7 mg/kg and 546.7 mg/kg, respectively. The suppression of orienting responses in the animals was noted when it was administered in neurotoxic doses. Valprazolamide showed the most antiepileptic activity on models of MES, scPTZ and isoniazid antagonism tests. The ED50 values were 138.4 mg/kg, 74.5 mg/kg, and 126.8 mg/kg, respectively. The therapeutic indices for these models of epilepsy were 6.7; 12.4; 7.3, and protective index – 3.3; 6.1 and 3.6, respectively. In the hot plate test, valprazolamide increased the latency period before a defensive response to a thermal stimulus (ED50 165 mg/kg). Conclusion: N-(5-ethyl-1,3,4-thiadiazol-2-yl)-2-propylpentanamide is a new 1,3,4-thiadiazolylamide derivative of 2-propylpentanoic acid with antiepileptic and analgesic activities, which belongs to the group of low-toxic agents. Graphic abstract N-(5-ethyl-1,3,4-thiadiazol-2-yl)-2-propylpentanamide (3D) LD50=924.8 mg/kg (mice, intraperitoneally) TD50=456.7 mg/kg (rotarod, mice, intraperitoneally) ED50=138.4 mg/kg (MES, mice, intraperitoneally) ED50=74.5 mg/kg (scPTZ, mice, intraperitoneally)

Effects of Folinic Acid Administration on Lower Limb Ischemia/Reperfusion Injury in Rats

Surgery under ischemic conditions, lasting up to 3 h, is routinely performed in orthopedic surgery, causing undesirable injury due to ischemia-reperfusion syndrome, with short and medium-term functional repercussions. To date, there is no established prophylactic treatment. In this work we evaluated folinic acid (FA) in a rodent model of lower limb ischemia-reperfusion (IRI-LL). 36 male WAG rats underwent 3 h of lower limb ischemia. In the saline group, rats received intraperitoneal administration of saline (used as vehicle for treatment). In the experimental group, rats were pretreated with FA (2.5 mg/kg) before the end of ischemia. After ischemia, animals were sacrificed at 3 h, 24 h or 14 days (for biochemical determination (Na+, K+, Cl-, urea, creatinine, CK, LDH, ALP, ALT, and AST), pathological assessment, or functional study using the rotarod test; respectively). Another six animals were used to establish the reference values. The prophylactic administration of FA significantly reduced the elevation of biochemical markers, especially those that most directly indicate muscle damage (CK and LDH). In addition, it also improved direct tissue damage, both in terms of edema, weight, PMN infiltrate and percentage of damaged fibers. Finally, the administration of FA allowed the animals to equal baseline values in the rotarod test; what did not occur in the saline group, where pre-ischemia levels were not recovered. Following 3 h of lower limb ischemia, FA minimizes the increase of CK and LDH, as well as local edema and leukocyte infiltration, allowing a faster recovery of limb functionality. Therefore, it could be considered as a prophylactic treatment when tourniquet is used in clinics.

The anxiolytic and antidepressant effect of Buxus hyrcana in the pentylenetetrazole kindled rat

Pentylenetetrazole (PTZ) is a chemical substance which largely used for induction of seizure and epilepsy in the animal model, and it can also, disrupts free radicals balance and causes oxidative stress in the body with a negative impact on behavioral statuses like anxiety and depression. In this study, the medicinal plant Buxus hyrcana, was used to evaluate its effect on oxidative stress, anxiety and depression caused by PTZ in the rat. Twenty-four male rats were randomly allocated to 4 groups: control negative under treatment with PTZ (sub-threshold dose 35 mg/kg for one month), control positive under treatment with phenobarbital (PB-30 mg/kg), and two PTZ groups under treatment with B. hyrcana extract (BHE-300, and -600 mg/kg). For anxiety parameters, the elevated plus maze (EPM) was used. The forced swim test (FST) and rotarod test were employed to assess the antidepressant and balance potential, respectively. After behavioral evaluation, rats were anesthetized, brains were removed, and following preparation of brain homogenates, oxidative stress was evaluated using specified methods. BHE administered at the doses of 300, and 600 mg/kg, reduced immobility time in the FST exerting antidepressant-like activity. In the EPM test, BHE at the same doses, produced the anxiolytic-like effect. Also, the rats which received BHE had a significant improvement in rotarod test in contrast to control groups. In addition, brain catalase activity and superoxide dismutase level were significantly greater versus PTZ group BHE-300 treated PTZ group was significantly lower and. BHE could prevent anxiety and depression and ameliorate oxidative stress in PTZ-kindled rats.

Analgesic Effect of Boiogito, a Japanese Traditional Kampo Medicine, on Post-Traumatic Knee Osteoarthritis through Inhibition of ERK1/2 Phosphorylation in the Dorsal Horn of the Spinal Cord

Boiogito (BO), a Japanese traditional herbal medicine, has been proven to be clinically effective against knee osteoarthritis (KOA)-associated pain. However, the therapeutic mechanism of BO remains unclear. Thus, we investigated the analgesic mechanism of BO using a rat KOA model. KOA was induced by destabilization of the medial meniscus (DMM). Rats were allocated into the following four groups: control, sham, DMM, and DMM + BO groups. Rotarod test was performed to evaluate the pain-related locomotive dysfunction. Expression of phosphorylated extracellular signal-regulated kinase1/2 (pERK1/2) in the spinal dorsal horn was examined using immunofluorescence staining and Western blotting on days 1 and 28 after DMM surgery. A mitogen-activated protein kinase inhibitor, U0126, was intrathecally injected and rotarod test and Western blotting were performed. The rotarod test revealed hampered locomotive function in the DMM group, which was significantly improved upon BO administration. The number of pERK1/2-positive cells was increased in the DMM group, whereas it was significantly decreased in the DMM + BO group. U0126 significantly inhibited ERK1/2 phosphorylation and increased walking time in the rotarod test, suggesting that the DMM-related pain was associated with ERK1/2 phosphorylation in the spinal dorsal horn. In conclusion, BO administration improved the pain-related locomotive dysfunction by suppressing ERK1/2 phosphorylation.

Synthesis, Spectral Characterization and Anticonvulsant Studies of the Novel Triazolothiadiazoles Bearing Benzoxazole Moiety

In this study, new fused triazolo-thiadiazoles (4a-o) were synthesized viamethyl 2-[bromo(phenyl)methyl]-1,3-benzoxazole-5-carboxylate. The structure of novel derivatives was recognized on the basis of spectral data results and screened their anticonvulsant action by means of maximal electroshock seizure (MES) and subcutaneous pentylenetetrazol (scPTZ) procedures. Minimal motor studies were completed by a rotarod method. Compounds 4e, 4g, 4j, 4l, 4m and 4n showing better anticonvulsant action corresponding to hydrophobicity. Other molecules remained fewer lipophilic and have less effectiveness. Most of the compounds positively tolerable the rotarod test deprived of motor deficiency. In conclusion, the prepared derivatives with distal aryl moiety exhibited higher lipophilic character and lead to improved pharmacological achievement, which can be a forthcoming promise.

Delta Sleep-Inducing Peptide Recovers Motor Function in SD Rats after Focal Stroke

Background and Objectives: Mutual effect of the preliminary and therapeutic intranasal treatment of SD rats with DSIP (8 days) on the outcome of focal stroke, induced with intraluminal middle cerebral occlusion (MCAO), was investigated. Materials and Methods: The groups were the following: MCAO + vehicle, MCAO + DSIP, and SHAM-operated. DSIP or vehicle was applied nasally 60 (±15) minutes prior to the occlusion and for 7 days after reperfusion at dose 120 µg/kg. The battery of behavioral tests was performed on 1, 3, 7, 14, and 21 days after MCAO. Motor coordination and balance and bilateral asymmetry were tested. At the end of the study, animals were euthanized, and their brains were perfused, serial cryoslices were made, and infarction volume in them was calculated. Results: Although brain infarction in DSIP-treated animals was smaller than in vehicle-treated animals, the difference was not significant. However, motor performance in the rotarod test significantly recovered in DSIP-treated animals. Conclusions: Intranasal administration of DSIP in the course of 8 days leads to accelerated recovery of motor functions.

Protective Effect of Monoisoamyl-2, 3-Dimercaptosuccinic Acid against Manganese-induced Neurotoxicity in Rats

Background: Apart from being an essential heavy metal, Manganese (Mn) serves as an important component of the antioxidant enzyme system in humans. Overexposure to manganese leads to the development of manganism, which is characterized by motor dysfunction along with neurodegeneration. The management of manganism often utilizes chelation therapy. In this regard, Monoisoamyl-2, 3-Dimercaptosuccinic Acid (MiADMSA) has been reported as a novel arsenic chelator, due to the presence of vicinal sulfhydril group. MiADMSA has been reported to reduce the level in divalent ions (like copper) therefore, it may be hypothesized that MiADMSA would be helpful in Mn-induced neurotoxicity. Objective: This study is envisaged to explore the protective effect of MiADMSA on Mn-induced neurotoxicity. Method: Mn exposure was carried out by intraperitoneal administration of Mn (as manganese chloride, 10 mg/kg; i.p.). The animals were treated with MiADMSA (50 mg/kg; p.o.) either alone or in combination with Mn. The effect of different treatments on neurobehavioral functions was observed by assessing spontaneous locomotor activity, motor rotarod test, and depression-like behavior in the forced swim test. After behavioral evaluations, all the animals were sacrificed and the brain and liver were isolated for metal estimations. Results: Mn exposure leads to loss of motor coordination as observed in spontaneous locomotor activity and rotarod test. However, treatment with MiADMSA significantly improved motor impairments as compared to Mn exposed animals. Accumulation of Mn in the liver and brain has been recorded with Mn exposure; however, MiADMSA treatment significantly reduced the Mn content from the liver and brain. Conclusion: The outcome of the study suggests that treatment with MiADMSA reversed Mn-induced neurotoxicity by reducing Mn load. Therefore, the use of MiADMSA may be suggested in manganese toxicity, after careful investigation.

Investigation of the Combination of Pregabalin with Duloxetine or Amitriptyline on the Pharmacokinetics and Antiallodynic Effect During Neuropathic Pain in Rats

BACKGROUND: Amitriptyline, duloxetine, and pregabalin are among the most pharmacotherapeutic, effective treatments for neuropathic pain control. However, the evaluation of synergism by combining these treatments is still poorly investigated. OBJECTIVES: To evaluate the pharmacokinetics of the combination of pregabalin plus duloxetine and pregabalin plus amitriptyline, as well as the effect of these on neuropathic pain on rodent model. STUDY DESIGN: The experimental study. SETTING: The research took place in the research laboratories at the Federal University of Alfenas after ethics committee approval. METHODS: This study used male Wistar rats weighing between 220 and 250 g. The animals were randomly divided into the following groups: monotherapy (pregabalin, amitriptyline, duloxetine), combined therapy (pregabalin + amitriptyline, pregabalin + duloxetine), and vehicle (ultrapure water). Pharmacokinetic analysis of pregabalin or combination (pregabalin + amitriptyline or pregabalin + duloxetine) in the plasma were performed by ultraperformance liquid chromatography tandem mass spectrometry. Neuropathic pain was induced by sciatic nerve constriction (chronic constriction injury [CCI]) model, and nociceptive threshold was measured by von Frey filaments test. In addition, to evaluate the influence of the treatments on the motor coordination, the rotarod test was used. RESULTS: The pharmacokinetic disposition of pregabalin was changed in the association with amitriptyline, presenting a clearance reduction and consequently an increase in bioavailability. Furthermore, after the 14th day of CCI, pregabalin was administered orally and induced antiallodynic effect after 1, 2:15, 4, and 8 hours of its administration and showed the greatest antiallodynic effect after 4 hours of its administration. Moreover, this effect was prolonged (up to 8 hours) by combination with amitriptyline. Additionally, pregabalin and pregabalin + duloxetine showed a hypoalgesic effect in sham rats. In addition, the rotarod test results showed that drugs did not influence the motor coordination of the rats. LIMITATIONS: Potential competition mechanisms during the excretion of pregabalin, when pregabalin was combined with amitriptyline, were not investigated in this study. CONCLUSIONS: The data demonstrated that combined therapy of pregabalin plus amitriptyline improved the bioavailability of pregabalin and potentiated the efficacy of the antiallodynic effect of pregabalin alone, proving to be advantageous for the treatment of sciatic neuropathic pain. KEY WORDS: Neuropathic pain, pregabalin, duloxetine, amitriptyline, pharmacokinetic, antiallodynic effect, combined therapy, rats

Antiparkinsonian Activity of Aqueous Extract of Agaricus Blazei Murill in Rotenone-induced Parkinson’s Disease

Background: Parkinsons disease is a chronic neurological disorder which may be due to reduction in the dopaminergic neurons in the brain. However, Agaricus blazei is a rich source of natural antioxidants. Aim: In this study, antiparkinsonian activity of Agaricus blazei Murill was evaluated using different animal models. Method: Antiparkinsonian activity was evaluated using two different doses (273 mg/kg and 819 mg/kg) of Agaricus blazei Murill. Rotenone and sunflower oil were used as positive and negative control, respectively. Catalepsy test, rotarod test, exploratory behavior test (rearing) and locomotor activity test were conducted to observe antiparkinsonian activity of the drug in rats. Result: The results of the animal models were confirmed by determining the levels of reduced glutathione, total protein, thiobarbituric acid reactive substances (TBARS) and nitric oxide in the animal brain. Pretreatment with Agaricus blazei Murill, showed marked reduction in rotenone-induced catalepsy and a significant increase in exploratory behavior, muscular activity, and locomotor activity in rats. Agaricus blazei Murill has also shown extremely significant effect in decreasing the oxidative stress in the animal brain by increasing the brain levels of reduced GSH and total proteins and decreasing the levels of nitrite and TBARS. Conclusion: The results of rotenone-induced catalepsy, exploratory behavior, rotarod test and locomotor activity showed that Agaricus blazei Murill exerts a significant ameliorative effect on Parkinson’s disease in rats.

PROFILATIC TREATMENT WITH FOLIC ACID FOR LOWER LIMB ISCHAEMIA-REPERFUSION INJURY IN AGED RATS

INTRODUCTION Lower limb ischemia-reperfusion injury (IRI-LL) is a common major complication in orthopaedic surgery, especially in elderly patients. We previously demonstrated that folic acid reduced IRI-LL damage in 3–4-month-old rats. Current work analyses the effect of folic acid in the prevention of IRI-LL in elderly animals. MATERIAL AND METHODS Thirty-six 12-month-old male WAG/RijHsd rats were subjected to 3 hours of ischaemia. Eighteen animals received folic acid (2.5 mg/kg, ip) 20 minutes before the end of ischaemia period; the other half received the same volume of saline. Animals were sacrificed after 3 hours, 24 hours, and 14 days of reperfusion for biochemical (tissue damage markers and electrolytes), histopathological studies of gastrocnemius muscle, and daily assessment of limb function by Rotarod test, respectively. RESULTS The administration of folic acid prior to the end of ischaemia period reduced by 30-40% the increase in LDH and CK observed in non-treated animals (p < 0.0001). When histological sections were analysed, folic acid was found to reduce by 20% the number of damaged muscle fibbers per field (60±17.1 vs 80.7±16.4, p < 0.0001). Rotarod test revealed that folic acid also led to an improvement in muscle function, assessed by the time the animals spend running on the rod, when comparing to untreated animals. CONCLUSIONS The administration of folic acid, prior to the end of ischemic period, decreases the damaged induced by IRI-LL, also achieving a faster recovery of mobility.