Long-term treatment with supraphysiological doses of nandrolone decanoate reduces the sensitivity of Bezold–Jarisch reflex control of heart rate and blood pressure

We investigated the influence of long-term treatment with supraphysiological doses of an anabolic-androgenic steroid on the Bezold–Jarisch reflex (BJR) control of heart rate (HR) and diastolic arterial pressure (DAP), and whether this treatment induced cardiac hypertrophy. Male rats were treated with nandrolone decanoate (ND) (10 mg kg⁻¹ body weight for 8 weeks; DECA) or vehicle (control animals; CON). After 8 weeks of treatment, the BJR was evaluated by bradycardia and hypotension responses that were elicited by serotonin administration (2–32 μg kg⁻¹). Mean arterial pressure (MAP) was assessed and cardiac hypertrophy was determined by the ratio of the left and right ventricle weight/body weight (LVW/BW and RVW/BW, respectively) and by histological analysis. Total body protein (TBP) content was also evaluated. Nandrolone decanoate treatment increased MAP (CON = 99 ± 1 mmHg; DECA = 109 ± 2 mmHg; p < 0.01) but did not change the mean basal HR (CON = 356 ± 13 bpm; DECA = 367 ± 11 bpm). The treatment also induced LV and RV hypertrophy (LVW/BW: CON = 1.86 ± 0.04 mg g⁻¹, DECA = 2.17 ± 0.04 mg g⁻¹, p < 0.01; RVW/BW: CON = 0.42 ± 0.02 mg g⁻¹, DECA = 0.53 ± 0.03 mg g⁻¹, p < 0.05) and reduced the number of myocyte nuclei/high-power field (CON = 23.0 ± 2; DECA = 9.4 ± 1.0; p < 0.01). ND treatment blunted the HR and DAP decreases induced by serotonin. ND determines an increase in the TBP content in DECA group (35 ± 3%; p < 0.01) compared with control animals (18 ± 1%). We conclude that 8 weeks of ND treatment induces anabolic effect, cardiac hypertrophy and an elevation of MAP. This treatment also reduces the sensitivity of the BJR control of bradycardia and blood pressure, possibly due to cardiac hypertrophy. The blunted BJR response could contribute to the MAP elevation in DECA animals.     

Toxicokinetics of glyphosate and its metabolite aminomethyl phosphonic acid in rats

The toxicokinetics of glyphosate after single 100 mg kg⁻¹ intravenous (i.v.) and 400 mg kg⁻¹ oral doses were studied in rats. Serial blood samples were obtained after i.v. and oral administration. Plasma concentrations of glyphosate and its metabolite amiomethyl phosphonic acid (AMPA) were determined by HPLC method. After i.v. and oral administration, plasma concentration–time curves were best described by a two-compartment open model. For glyphosate, the elimination half-lives (T_1/2β) from plasma were 9.99 h after i.v. and 14.38 h after oral administration. The total plasma clearance was not influenced by dose concentration or route and reached a value of 0.995 l h⁻¹ kg⁻¹. After i.v. administration, the apparent volume of distribution in the second compartment (V₂) and volume of distribution at steady state (V_ss) were 2.39 and 2.99 l kg⁻¹, respectively, suggesting a considerable diffusion of the herbicide into tissues. After oral administration, glyphosate was partially and slowly absorbed with a T_max of 5.16 h. The oral bioavailability of glyphosate was found to be 23.21%. Glyphosate was converted to AMPA. The metabolite AMPA represented 6.49% of the parent drug plasma concentrations. The maximum plasma concentrations of glyphosate and AMPA were 4.62 and 0.416 μg ml⁻¹, respectively. The maximum plasma concentration of AMPA was achieved at 2.42 h. For AMPA, the elimination half-life (T_1/2β) was 15.08 h after oral administration of glyphosate parent compound.     

Benfotiamine attenuates nicotine and uric acid-induced vascular endothelial dysfunction in the rat

The study has been designed to investigate the effect of benfotiamine, a thiamine derivative, in nicotine and uric acid-induced vascular endothelial dysfunction (VED) in rats. Nicotine (2 mg kg⁻¹ day⁻¹, i.p., 4 weeks) and uric acid (150 mg kg⁻¹ day⁻¹, i.p., 3 weeks) were administered to produce VED in rats. The development of VED was assessed by employing isolated aortic ring preparation and estimating serum and aortic concentration of nitrite/nitrate. Further, the integrity of vascular endothelium was assessed using the scanning electron microscopy (SEM) of thoracic aorta. Moreover, the oxidative stress was assessed by estimating serum thiobarbituric acid reactive substances (TBARS) and aortic superoxide anion generation. The administration of nicotine and uric acid produced VED by impairing the integrity of vascular endothelium and subsequently decreasing serum and aortic concentration of nitrite/nitrate and attenuating acetylcholine-induced endothelium dependent relaxation. Further, nicotine and uric acid produced oxidative stress, which was assessed in terms of increase in serum TBARS and aortic superoxide generation. However, treatment with benfotiamine (70 mg kg⁻¹ day⁻¹, p.o.) or atorvastatin (30 mg kg⁻¹ day⁻¹ p.o., a standard agent) markedly prevented nicotine and uric acid-induced VED and oxidative stress by improving the integrity of vascular endothelium, increasing the concentration of serum and aortic nitrite/nitrate, enhancing the acetylcholine-induced endothelium dependent relaxation and decreasing serum TBARS and aortic superoxide anion generation. Thus, it may be concluded that benfotiamine reduces the oxidative stress and consequently improves the integrity of vascular endothelium and enhances the generation of nitric oxide to prevent nicotine and uric acid-induced experimental VED.     

Comparison of verapamil and bepridil,two slow channel inhibitors,in protection against calcium-induced arrhythmias

Summary Verapamil and bepridil share the common property of antagonizing the slow inward calcium-mediated current, but bepridil has some additional antiarrhythmic properties. The efficacy of these two compounds against CaCl₂-induced arrhythmias has been compared in rats. CaCl₂ was administered i.v. by continuous infusion until death (25 mg·kg^–1·min^–1 or 40 mg·kg^–1·min^–1) or by bolus injection (160 mg·kg^–1). Bepridil (5, 10 mg·kg^–1) or verapamil (2.5,5 mg·kg^–1) were injected 10 min before CaCl₂. Bepridil (10 mg·kg^–1) or verapamil (5 mg·kg^–1) prolong the survival time during CaCl₂ infusion. After pretreatment, the injection of 160 mg·kg^–1 CaCl₂ is less toxic: 25% of animals are protected by bepridil (5 mg·kg^–1), 41% by bepridil (10 mg·kg^–1) or verapamil (5 mg·kg^–1).At death the myocardial Ca²⁺ level is not different in controls and pretreated animals, thus, the ratio myocardial Ca²⁺/total injected Ca²⁺ is significantly lowered by bepridil (10 mg·kg^–1) or verapamil (5 mg·kg^–1). The efficacy of the two drugs on this model appears related solely to inhibition of slow inward current despite the additional antiarrhythmic profile of bepridil.     

Pharmacokinetics of thrombin-like enzyme from venom of Agkistrodon halys ussuriensis Emelianov determined by ELISA in the rat

A thrombin-like enzyme (TLE) was separated and purified from the venom of a northeast Chinese snake Agkistrodon halys ussuriensis Emelianov. Experiments were performed in rats to determine the pharmacokinetic parameters following an intravenous (i.v.) or a subcutaneous (s.c.) injection of the thrombin-like enzyme. The plasma levels of TLE were estimated by enzyme-linked immunosorbent assay. The method exhibited high reproducibility and accuracy in correlating optical densities with TLE concentrations (0.2–30 ng ml⁻¹, r=0.99). The plasma concentration-time course after i.v. administration of 50 μg kg⁻¹ TLE was well fitted by a two-compartment open model. The half-life of the α-phase was 18.0±3.2 min, and that of the β-phase 3.9±0.7 h. The apparent volume of distribution was 1.8±0.5 l kg⁻¹, and clearance was 5.4±0.5 ml min⁻¹ kg⁻¹. When the TLE was injected s.c. at a dose of 0.75 mg kg⁻¹, the changes in plasma concentration were best described by a two-compartment model with a first-order absorption. The maximal plasma level of 51±2.7 ng ml⁻¹ was reached at 5.2±0.5 h. The absorption rate constant was 0.3±0.03 h⁻¹. The area under the plasma concentration-time curve (AUC) was 2.8±0.8 μg h⁻¹ ml⁻¹.     

Phyllaemblicin B inhibits Coxsackie virus B3 induced apoptosis and myocarditis

Coxsackie virus B3 (CVB3) is believed to be a major contributor to viral myocarditis since virus-associated apoptosis plays a role in the pathogenesis of experimental myocarditis. In this study, we investigated the in vitro and in vivo antiviral activities of Phyllaemblicin B, the main ellagitannin compound isolated from Phyllanthus emblica, a Chinese herb medicine, against CVB3. Herein we report that Phyllaemblicin B inhibited CVB3-mediated cytopathic effects on HeLa cells with an IC₅₀ value of 7.75 ± 0.15 μg/mL. In an in vivo assay, treatment with 12 mg kg⁻¹ d⁻¹ Phyllaemblicin B reduced cardiac CVB3 titers, decreased the activities of LDH and CK in murine serum, and alleviated pathological damages of cardiac muscle in myocarditic mice. Moreover, Phyllaemblicin B clearly inhibited CVB3-associated apoptosis effects both in vitro and in vivo. These results show that Phyllaemblicin B exerts significant antiviral activities against CVB3. Therefore, Phyllaemblicin B may represent a potential therapeutic agent for viral myocarditis.     

A comparison of the stability of doxorubicin and daunorubicin in solid state

The degradation of doxorubicin and daunorubcin in the solid state was studied using an HPLC method with UV detection (LiChrospher RP-18, 5 μm, 250 mm × 4 mm; mobile phase: acetonitrile-solution A 1:1, v/v (solution A: 2.88 g of laurisulfate sodium and 1.6 ml of phosphoric acid(V) in 1000 ml); flow rate – 1.4 ml min⁻¹; UV detection – 254 nm). The degradation of doxorubicin was a first-order reaction depending on the substrate concentration and daunorubicin degraded according to the kinetic model of autocatalysis. The dependence ln k_i = f(1/T) was described by the equations ln k_DOX = 40.0 ± 15.6 – (19804 ± 5682) (1/T) and ln k_DAU = 35.9 ± 11.3 – (16581 ± 3972) (1/T) at 76.4% RH. The dependence ln k_i = f(RH%) was described by the equations ln k_DOX = (8.80 ± 3.60) × 10⁻² (RH%) – (21.50 ± 2.57) and ln k_DAU = (6.63 ± 1.22) × 10⁻² (RH%) – (13.35 ± 1.68). The thermodynamic parameters (E_a, ΔH^≠a, ΔS^≠a) of the degradation of doxorubicin and daunorubicin were calculated. Although the degradation of doxorubicin was slower at increased temperature (353–373 K) and relative air humidity (50.9–90.0%), the differences between the influence of temperature and relative air humidity on the stability doxorubicin and of daunorubicin were not significant.     

Reduced number of α- and β-adrenergic receptors in the myocardium of rats exposed to tobacco smoke

The concentration of α- and β-adrenergic receptors-as measured by specific [³H]WB-4101 and (−)-[³H]-dihydroalprenolol binding-was diminished by 60% below control values in the hearts of hearts of rats exposed to tobacco smoke. These changes in receptor numbers took place almost immediately after tobacco smoke exposure and were rapidly reversible after termination of the exposure. The dissociation constant, K , for [³H]WB-4101 was identical in exposed (K_D = 0.34 ± 0.09 nM) and control (K_D = 0.35 ± 0.07 nM) hearts but was significantly different i in the case of (-)-[³H] dihydroalprenolol binding (exposed, (K_D = 2.83 ± 0.30 nM vs control K_D = 5.22 ± 0.61 nM). For β-receptor binding there was no significant difference between exposed and control animals in the K_i values (−)-epinephrine, (−)-norepinephrine, (−)-alprenolol, (±)-propranolol or timolol. (−)-Isoproterenol, however, was found to bind with lower affinity in exposed compared with control hearts. For α-receptor binding there was no significant difference between control and ‘smoked’ animals in the K_i values for (−)-epinephrine, (−)-norepinephrine or phentolamine. The decrease in α- and β-adrenergic receptor concentration may be related to the phenomenon of receptor desensitization resulting from a release of catecholamines in rats exposed to tobacco smoke.     

Citalopram-induced hypophagia is enhanced by blockade of 5-HT1A receptors: role of 5-HT2C receptors

1. The selective 5-hydroxytryptamine reuptake inhibitor citalopram (10 and 20 mg kg⁻¹, i.p.) significantly reduced food intake in male rats (CD-COBS) habituated to eat their daily food during a 4-h period.
2. The 5-HT_1A receptor antagonist WAY100635 (0.3 mg kg⁻¹) administered systemically did not modify feeding but significantly potentiated the reduction in food intake caused by 10 mg kg⁻¹ i.p. citalopram. The dose of 5 mg kg⁻¹ i.p. citalopram was not active in animals pretreated with vehicle but significantly reduced feeding in animals pretreated with WAY100635.
3. WAY100635 (0.1 μg 0.5 μl⁻¹) injected into the dorsal raphe significantly potentiated the hypophagic effect of 10 mg kg⁻¹ citalopram.
4. WAY100635 (1.0 μg 0.5 μl⁻¹) injected into the median raphe did not modify feeding or the hypophagic effect of 10 mg kg⁻¹ citalopram.
5. The 5-HT_2B/2C receptor antagonist SB206553 (10 mg kg⁻¹, p.o.) slightly reduced feeding by itself but partially antagonized the effect of WAY100635 administered systemically (0.3 mg kg⁻¹, s.c.) or into the dorsal raphe (0.1 μg 0.5 μl⁻¹) in combination with 10 mg kg⁻¹ i.p. citalopram. The hypophagic effect of 10 mg kg⁻¹ i.p. citalopram alone was not significantly modified by SB206553.
6. Brain concentrations of citalopram and its metabolite desmethylcitalopram in rats pretreated with SB206553, WAY100635 and their combination were comparable to those of vehicle-pretreated rats, 90 min after citalopram injection.
7. The hypophagic effect of citalopram was potentiated by blocking 5-HT_1A receptors. Only the effect of the WAY100635/citalopram combination seemed to be partially mediated by central 5-HT_2C receptors.

     

Embryo–fetal toxicity assessment of vonoprazan in rats and rabbits

Vonoprazan is a new potassium‐competitive acid blocker to treat acid‐related diseases. However, its safety during pregnancy is unclear. The aim of the study was to investigate the potential reproductive toxicity on the embryo–fetal development of vonoprazan. Vonoprazan acetate was administered by intravenous injection to pregnant rats (0, 2, 6 and 20 mg kg^–1 day^–1) and rabbits (0, 1.2, 3.6 and 12 mg kg^–1 day^–1) during the organogenetic period (gestation day 6–15 [rats] and 6–18 [rabbits]). Maternal reproductive endpoints were evaluated, together with effects on fetal growth and morphological development. In rats, no treatment‐related effects were found in the highest dose group (20 mg kg^–1) and the maternal plasma exposure was ≥50‐fold the expected clinical human exposure. However, in rabbits, dose‐related clinical signs (soft or liquid feces) occurred in the 12 mg kg^–1 group, which was regarded as a maternal toxicity. Besides, decreased maternal weight gain also was considered as a minimal maternal toxicity. At 12 mg kg^–1, delayed fetal ossification was found as evidence of embryo–fetal growth retardation, which was related to decreased fetal and placental weights. There was no maternal and developmental toxicity in the 1.2 and 3.6 mg kg^–1 groups. Thus, the no‐observed‐adverse‐effect levels of vonoprazan acetate in rabbits are considered 3.6 mg kg^–1 day^–1, which produced plasma exposure that was about 18‐fold human clinical exposure.     

Occurrence of Deoxynivalenol in Wheat in Slovakia during 2010 and 2011

In this study, a total of 299 grain samples of wheat were collected from four production regions: the maize, sugar beet, potato and feed sectors of Slovakia. The samples were analyzed for deoxynivalenol (DON) content by using an enzyme-linked immunosorbent assay Ridascreen^® Fast DON. Analysis of variance revealed a significant difference between years in DON contents (p < 0.027). The occurrence of samples with DON was 82.2% in 2010, with maximum DON content of 7.88 mg kg⁻¹, and 70.7% in 2011, with maximum DON content of 2.12 mg·kg⁻¹. The total mean DON content was 0.62 mg·kg⁻¹; in the feed region 0.22 mg·kg⁻¹; 0.63 mg·kg⁻¹ in the maize region; 0.78 mg·kg⁻¹ in the sugar beet region; 0.45 mg·kg⁻¹ the potato region. The limit of 1.25 mg·kg⁻¹ imposed by the European Union (EU) for DON content was exceeded in 13.7% of the studied samples. The average monthly rainfall for May to June played a critical role in DON content of wheat grains for maize and sugar beet producing regions. The present results indicate that DON content was at a high level in grains from wheat grown during 2010.     

α-Monofluoromethyldopa (MFMD) in combination withl-DOPA

Summary We have investigated the interaction of -monofluoromethyldopa (MFMD) with the effects of i.p. injectedl-DOPA (200 mg·kg^–1) on blood pressure and tissue catecholamines in normal and spontaneously hypertensive rats (SHR). MFMD 10 mg·kg^–1 (i.p.) effectively antagonizes thel-DOPA induced increase in heart dopamine (DA). This action is also seen after 15 or 50 mg·kg^–1. The accumulation of DA in the brain is very much reduced by MFMD 50 mg ·kg^–1 while after 15 or, especially, 10 mg·kg^–1 more DA is formed in the rrain than afterl-DOPA alone, probably due to the peripheral decarboxylase inhibition which presents morel-DOPA to the brain. We conclude that MFMD 10 mg ·kg^–1 gives a relatively selective peripheral inhibition of the decarboxylation ofl-DOPA and this dose combination was accordingly found to result in a reduction of blood pressure in conscious animals. This hypotensive response tol-DOPA was attenuated after MFMD 15 mg·kg^–1 and was absent after MFMD 50 mg·kg^–1. Interestingly, the hypotensive effect ofl-DOPA after MFMD 10 or 15 mg·kg^–1 was more pronounced in SHR.     

Effects of inhibitors of the activity of poly (ADP-ribose) synthetase on the liver injury caused by ischaemia-reperfusion: a comparison with radical scavengers

1. Poly (ADP-ribose) synthetase (PARS) is a nuclear enzyme activated by strand breaks in DNA which are caused by reactive oxygen species (ROS) and peroxynitrite. Excessive activation of PARS may contribute to the hepatocyte injury caused by ROS in vitro and inhibitors of PARS activity reduce the degree of reperfusion injury of the heart, skeletal muscle and brain in vivo. Here we compared the effects of various inhibitors of the activity of PARS with those of deferoxamine (an iron chelator which prevents the generation of hydroxyl radicals) and tiron (an intracellular scavenger of superoxide anion) on the degree of hepatic injury caused by ischaemia and reperfusion of the liver in the anaesthetized rat or rabbit.
2. In the rat, ischaemia (30 or 60 min) and reperfusion (120 min) of the liver resulted in significant increases in the serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) indicating the development of liver injury. Intravenous administration of the PARS inhibitors 3-aminobenzamide (3-AB, 10 mg kg⁻¹ or 30 mg kg⁻¹), 1,5-dihydroxyisoquinoline (ISO, 1 mg kg⁻¹) or 4-amino-1,8-naphthalimide (4-AN, 3 mg kg⁻¹) before reperfusion did not reduce the degree of liver injury caused by ischaemia-reperfusion.
3. In contrast to the PARS inhibitors, deferoxamine (40 mg kg⁻¹) or tiron (300 mg kg⁻¹) significantly attenuated the rise in the serum levels of AST and ALT caused by ischaemia-reperfusion of the liver of the rat.
4. In the rabbit, the degree of liver injury caused by ischaemia (60 min) and reperfusion (120 min) was also not affected by 3-AB (10 mg kg⁻¹) or ISO (1 mg kg⁻¹).
5. These results support the view that the generation of oxygen-derived free radicals mediates the liver injury associated with reperfusion of the ischaemic liver by mechanism(s) which are independent of the activation of PARS.

     

Antihypertensive and vasorelaxant activities of Laelia autumnalis are mainly through calcium channel blockade

The aim of the present study was to evaluate the possible mechanism of the vasorelaxant action of methanol extract from Laelia autumnalis (MELa) in isolated rat aortic rings, and to establish its antihypertensive activity in vivo. MELa (0.15→50 µg/mL) induced relaxation in aortic rings pre-contracted with KCl (80 mM), showing an IC₅₀ value of 34.61 ± 1.41 µg/mL and E_max value of 85.0 ± 4.38% (in endothelium-intact rings) and an IC₅₀ value of 45.11 ± 4.17 µg/mL and E_max value of 80.0 ± 12.1% (in endothelium-denuded rings). Serotonin (5-HT, 1 × 10^− 4 M) provoked sustained contraction, which was markedly inhibited by MELa (0.15→50 µg/mL) in a concentration-dependent and endothelium-independent manner. Pretreatment with MELa (15, 46, 150, 300 and 1500 µg/mL) also inhibited contractile responses to norepinephrine (NE 1 × 10^− 11 M to 1 × 10^− 5.5 M). In endothelium-denuded rings, the vasorelaxant effect of MELa was reduced partially by ODQ (1 µM), but not by tetraethylammonium (5 µM), glibenclamide (10 µM), and 2-aminopyridine (100 µM). The extract also reduced NE-induced transient contraction in Ca²⁺-free solution, and inhibited contraction induced by increasing external calcium in Ca²⁺-free medium plus high KCl (80 mM). The antihypertensive effect of MELa was determined in spontaneously hypertensive rats (SHR). A single oral administration of the extract (100 mg/kg) exhibited a significant decrease in systolic and diastolic blood pressure and heart rate (p < 0.05) in SHR rats. Our results suggest that MELa induces relaxation in rat aortic rings through an endothelium-independent pathway, involving blockade of Ca²⁺ channels and a possible cGMP enhanced concentrations and also causes an antihypertensive effect.     

A simplified screening procedure for determination of total N–NO groups (TNG) and nitrite (NO2) in commercial low-molecular-weight heparins (LMWH) by selective chemical denitrosation followed by high-sensitivity chemiluminescence detection (NO-analyzer, NOA)

Aim of this work was to set up a method for the sensitive and selective determination of nitrite (NO₂⁻) and total N-nitroso groups (TNG) in dalteparin and nadroparin, commercial low- molecular-weight heparins (LMWH), prepared by deaminative depolymerization of heparin with nitrous acid. The European Pharmacopoeia VI ed. indicates respectively 5 ppm as the maximum content for contaminant NO₂⁻ in the former and 0.25 ppm for TNG in the latter and no clear indication is given for N–NO groups in dalteparin, i.e. TNG must be absent because of the specific manufacturing process.The proposed technique is based on the development of a pre-analytical device, coupled to a chemiluminometer, constituted by three sequentially connected and commercially available purge vessels, where selective reagents are employed for the conversion of NO₂⁻ and N–NO to nitric oxide (NO). In detail, NO₂⁻ was determined in the first chamber and non-volatile and volatile TNG in the second and third. This method was validated for selectivity, sensitivity, linearity, accuracy and precision. The method was shown to be selective, with a quantitative linear range of 1–1000 ppb). The bias, intra- and inter-day percent relative error was lower than 1%. The contamination of NO₂⁻ and TNG in nadreparin was below the limits; for dalteparin NO₂⁻ fell within the limit, but there was a huge amount of TNG (15.80 ± 0.05 ppm–6.69 ± 0.02 ppm). Preliminary investigation on the solvent-extractable material from dalteparin showed the majority of chemiluminescence retained in the aqueous residue to indicate that this N–NO groups may belong to solvent unextractable material or be tightly bound to the dalteparin backbone.     

Gabapentin (neurontin) and S-(+)-3-isobutylgaba represent a novel class of selective antihyperalgesic agents

1. Gabapentin (neurontin) is a novel antiepileptic agent that binds to the α₂δ subunit of voltage-dependent calcium channels. The only other compound known to possess affinity for this recognition site is the (S)-(+)-enantiomer of 3-isobutylgaba. However, the corresponding (R)-(−)-enantiomer is 10 fold weaker. The present study evaluates the activity of gabapentin and the two enantiomers of 3-isobutylgaba in formalin and carrageenan-induced inflammatory pain models.
2. In the rat formalin test, S-(+)-3-isobutylgaba (1–100 mg kg⁻¹) and gabapentin (10–300 mg kg⁻¹) dose-dependently inhibited the late phase of the nociceptive response with respective minimum effective doses (MED) of 10 and 30 mg kg⁻¹, s.c. This antihyperalgesic action of gabapentin was insensitive to naloxone (0.1–10.0 mg kg⁻¹, s.c.). In contrast, the R-(−)-enantiomer of 3-isobutylgaba (1–100 mg kg⁻¹) produced a modest inhibition of the late phase at the highest dose of 100 mg kg⁻¹. However, none of the compounds showed any effect during the early phase of the response.
3. The s.c. administration of either S-(+)-3-isobutylgaba (1–30 mg kg⁻¹) or gabapentin (10–100 mg kg⁻¹), after the development of peak carrageenan-induced thermal hyperalgesia, dose-dependently antagonized the maintenance of this response with MED of 3 and 30 mg kg⁻¹, respectively. Similar administration of the two compounds also blocked maintenance of carrageenan-induced mechanical hyperalgesia with MED of 3 and 10 mg kg⁻¹, respectively. In contrast, R-(−)-3-isobutylgaba failed to show any effect in the two hyperalgesia models.
4. The intrathecal administration of gabapentin dose-dependently (1–100 μg/animal) blocked carrageenan-induced mechanical hyperalgesia. In contrast, administration of similar doses of gabapentin into the inflamed paw was ineffective at blocking this response.
5. Unlike morphine, the repeated administration of gabapentin (100 mg kg⁻¹ at start and culminating to 400 mg kg⁻¹) over 6 days did not lead to the induction of tolerance to its antihyperalgesic action in the formalin test. Furthermore, the morphine tolerance did not cross generalize to gabapentin. The s.c. administration of gabapentin (10–300 mg kg⁻¹), R-(−) (3–100 mg kg⁻¹) or S-(+)-3-isobutylgaba (3–100 mg kg⁻¹) failed to inhibit gastrointestinal motility, as measured by the charcoal meal test in the rat. Moreover, the three compounds (1–100 mg kg⁻¹, s.c.) did not generalize to the morphine discriminative stimulus. Gabapentin (30–300 mg kg⁻¹) and S-(+)-isobutylgaba (1–100 mg kg⁻¹) showed sedative/ataxic properties only at the highest dose tested in the rota-rod apparatus.
6. Gabapentin (30–300 mg kg⁻¹, s.c.) failed to show an antinociceptive action in transient pain models. It is concluded that gabapentin represents a novel class of antihyperalgesic agents.

     

Characterization of the heterozygous glucokinase knockout mouse as a translational disease model for glucose control in type 2 diabetes

Background and Purpose

The global heterozygous glucokinase (GK) knockout (gk^wt/del) male mouse, fed on a high-fat (60% by energy) diet, has provided a robust and reproducible model of hyperglycaemia. This model could be highly relevant to some facets of human type 2 diabetes (T2D). We aimed to investigate the ability of standard therapeutic agents to lower blood glucose at translational doses, and to explore the glucose-lowering potential of novel glucokinase activators (GKAs) in this model.

Experimental Approach

We measured the ability of insulin, metformin, glipizide, exendin-4 and sitagliptin, after acute or repeat dose administration, to lower free-feeding glucose levels in gk^wt/del mice. Further, we measured the ability of novel GKAs, GKA23, GKA71 and AZD6370 to control glucose either alone or in combination with some standard agents.

Key Results

A single dose of insulin (1 unit·kg⁻¹), metformin (150, 300 mg·kg⁻¹), glipizide (0.1, 0.3 mg·kg⁻¹), exendin-4 (2, 20 μg·kg⁻¹) and GKAs reduced free-feeding glucose levels. Sitagliptin (10 mg·kg⁻¹), metformin (300 mg·kg⁻¹) and AZD6370 (30, 400 mg·kg⁻¹) reduced glucose excursions on repeat dosing. At a supra-therapeutic dose (400 mg·kg⁻¹), AZD6370 also lowered basal levels of glucose without inducing hypoglycaemia.

Conclusion and Implications

Standard glucose-lowering therapeutic agents demonstrated significant acute glucose lowering in male gk^wt/del mice at doses corresponding to therapeutic free drug levels in man, suggesting the potential of these mice as a translatable model of human T2D. Novel GKAs also lowered glucose in this mouse model.     

Antidepressant-like effects of cannabidiol in mice: possible involvement of 5-HT1A receptors

Background and purpose:

Cannabidiol (CBD) is a non-psychotomimetic compound from Cannabis sativa that induces anxiolytic- and antipsychotic-like effects in animal models. Effects of CBD may be mediated by the activation of 5-HT_1A receptors. As 5-HT_1A receptor activation may induce antidepressant-like effects, the aim of this work was to test the hypothesis that CBD would have antidepressant-like activity in mice as assessed by the forced swimming test. We also investigated if these responses depended on the activation of 5-HT_1A receptors and on hippocampal expression of brain-derived neurotrophic factor (BDNF).

Experimental approach:

Male Swiss mice were given (i.p.) CBD (3, 10, 30, 100 mg·kg⁻¹), imipramine (30 mg·kg⁻¹) or vehicle and were submitted to the forced swimming test or to an open field arena, 30 min later. An additional group received WAY100635 (0.1 mg·kg⁻¹, i.p.), a 5-HT_1A receptor antagonist, before CBD (30 mg·kg⁻¹) and assessment by the forced swimming test. BDNF protein levels were measured in the hippocampus of another group of mice treated with CBD (30 mg·kg⁻¹) and submitted to the forced swimming test.

Key results:

CBD (30 mg·kg⁻¹) treatment reduced immobility time in the forced swimming test, as did the prototype antidepressant imipramine, without changing exploratory behaviour in the open field arena. WAY100635 pretreatment blocked CBD-induced effect in the forced swimming test. CBD (30 mg·kg⁻¹) treatment did not change hippocampal BDNF levels.

Conclusion and implications:

CBD induces antidepressant-like effects comparable to those of imipramine. These effects of CBD were probably mediated by activation of 5-HT_1A receptors.     

The role of α2-adrenoceptor antagonism in the anti-cataleptic properties of the atypical neuroleptic agent,clozapine, in the rat

1. The mechanism underlying the anticataleptic properties of the atypical neuroleptic agent, clozapine, has been investigated in the rat.
2. The close structural analogues of clozapine, loxapine (0.1 mg kg⁻¹ s.c.) and iso-clozapine (1 and 3 mg kg⁻¹ s.c.) induced catalepsy in rats. In contrast, clozapine and the regio-isomer of loxapine, iso-loxapine (up to 10 mg kg⁻¹ s.c.) did not produce catalepsy, but at a dose of 1 mg kg⁻¹ significantly inhibited catalepsy induced by loxapine (0.3 mg kg⁻¹ s.c.).
3. Radioligand binding assays showed that cataleptogenic potential was most clearly predicted by the D₂/5-HT_1A, D₂/5-HT_1B/1D and D₂/α₂-receptor affinity (K_D) ratios: i.e. 30–100-fold higher ratios were calculated for loxapine and iso-clozapine, whereas the ratios were less than 1 for clozapine and iso-loxapine. The ratios of affinities for D₂ to 5-HT_2A, 5-HT_2C or D₁ did not reflect the grouping of cataleptic and non-cataleptic compounds.
4. Co-treatment with the α₂-adrenoceptor antagonists, yohimbine (1–10 mg kg⁻¹ s.c.), RX 821002 (1–10 mg kg⁻¹ s.c.) and MK-912 (0.3 and 1 mg kg⁻¹ s.c.) dose-dependently inhibited the cataleptic response to loxapine (0.3 mg kg⁻¹). Yohimbine (1–10 mg kg⁻¹ s.c.) also dose-dependently inhibited the cateleptic response to haloperidol (0.3 mg kg⁻¹ s.c.). The α₂-adrenoceptor antagonists had no effect per se.
5. Neither yohimbine (10 mg kg⁻¹) nor RX821002 (3 mg kg⁻¹) altered the cataleptic response to the D₁ receptor antagonist, SCH 23390 (1 mg kg⁻¹ s.c.), while, like clozapine, both compounds abolished the response to the 5-HT_2A receptor antagonist, MDL 100,151 (3 mg kg⁻¹ s.c.).
6. The present data strongly implicate α₂-adrenoceptor blockade in the anticataleptic properties of clozapine and suggest that its lack of extrapyramidal side effects in the clinic may also be a consequence of this property.