S 47445 is a novel positive allosteric modulator of alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptors (AMPA-PAM). S 47445 enhanced glutamate's action at AMPA receptors on human and rat receptors and was inactive at NMDA and kainate receptors. Potentiation did not differ among the different AMPA receptors subtypes (GluA1/2/4 flip and flop variants) (EC50 between 2.5±5.4 μM), except a higher EC50 value for GluA4 flop (0.7 μM) and a greater amount of potentiation on GluA1 flop. A low concentration of S47445 (0.1 μM) decreased receptor response decay time of GluA1flop/GluA2flip AMPA receptors and increased the sensitivity to glutamate. Furthermore, S 47445 (0.1 and 0.3 μM) in presence of repetitive glutamate pulses induced a progressive potentiation of the glutamate-evoked currents from the second pulse of glutamate confirming a rapid-enhancing effect of S 47445 at low concentrations. The potentiating effect of S 47445 (1 μM) was concentration-dependently reversed by the selective AMPA receptor antagonist GYKI52466 demonstrating the selective modulatory effect of S 47445 on AMPA receptors. Using an AMPA-kainate chimera approach, it was confirmed that S 47445 binds to the common binding pocket of AMPA-PAMs. S 47445 did not demonstrate neurotoxic effect against glutamate-mediated excitotoxicity in vitro, in contrast significantly protected rat cortical neurons at 10 μM. S 47445 was shown to improve both episodic and spatial working memory in adult rodents at 0.3 mg/kg, as measured in the natural forgetting condition of object recognition and T-maze tasks. Finally, no deleterious effect on spontaneous locomotion and general behavior was observed up to 1000 mg/kg of S 47445 given acutely in rodents, neither occurrence of convulsion or tremors. Collectively, these results indicate that S 47445 is a potent and selective AMPA-PAM presenting procognitive and potential neuroprotective properties. This drug is currently evaluated in clinical phase 2 studies in Alzheimer's disease and in Major Depressive Disorder. download
Pharmacological characterisation of S 47445,a novel positive allosteric modulator of AMPAreceptors
A perspective on the contribution of animal models to the pharmacological treatment of posttraumatic stress disorder
Posttraumatic stress disorder (PTSD) is a prevalent, chronic, disabling disorder that may develop following exposure to a traumatic event. This review summarizes currently used animal models of PTSD and their potential role in the development of better therapeutics. Heterogeneity is one of the main characteristics of PTSD with the consequence that many pharmacological approaches are used to relieve symptoms of PTSD. To address the translational properties of the animal models, we discuss the types of stressors used, the rodent correlates of human PTSD (DSM-5) symptoms, and the efficacy of approved, recommended and off-label drugs used to treat PTSD in "PTSD-animals".
Methyllycaconitine- and scopolamine-induced cognitive dysfunction: differential reversal effect by cognition-enhancing drugs
There is a growing body of evidence pointing to the pivotal role of alpha-7 nicotinic acetylcholine receptor (α7 nAchR) dysfunction in cognitivedisorders such as Alzheimer's disease or schizophrenia. This study was undertaken to establish and characterize an in vivo model for cognitivedisorder secondary to the blockade of α7 nAChR by its specific antagonist, methyllycaconitine (MLA). The results show that MLA elicited cognitivedysfunction as assessed by reduced spontaneous alternation of mice in the T-maze. The maximal effect of MLA produced 25-30% reduction in the spontaneous alternation of mice, a level comparable with that induced by the muscarinic antagonism of scopolamine. Donepezil and galantamine fully reversed both MLA and scopolamine-induced cognitive dysfunction. However, the ED50 of donepezil and galantamine was significantly shifted to the left in the MLA- compared to scopolamine-treated mice (0.0005 and 0.002 mg/kg for donepezil; 0.0003 and 0.7 mg/kg for galantamine). Moreover, memantine elicited marked reversion of cognitive dysfunction (up to 70%) in MLA-treated mice while only a weak reversal effect at high dose of memantine (less than 20%) was observed in scopolamine-treated mice. The above findings indicate that MLA-induced cognitive dysfunction in the mouse is highly sensitive and more responsive to the current procognitive drugs than the traditional scopolamine-based assay. Thus, it can be of value for the preclinical screening and profiling of cognition-enhancing drugs. download
Treatment with Actovegin (R) Improves Sensory Nerve Function and Pathology in Streptozotocin-Diabetic Rats via Mechanisms Involving Inhibition of PARP Activation
BACKGROUND: Diabetic neuropathy is one of the most severe complications of diabetes, affecting approximately one-third of diabetic patients. We investigated the potential neuroprotective effect of Actovegin®, a deproteinized hemoderivative of calf blood, in an animal model of diabetic neuropathy.
METHODS :A single intravenous injection of streptozotocin (STZ, 55 mg/kg) was used to induce experimental diabetes in male Sprague-Dawley rats. Actovegin® (200 or 600 mg/kg) was administered intraperitoneally from day 11 to day 40 post-STZ exposure. N-acetylcysteine (NAC) was used as a positive control and was added to drinking water (0.2 g/l) from day 2 until day 40. Measurements to assess efficacy included sensory nerve conduction velocity (SNCV), intraepidermal nerve fiber density (IENFD), and poly(ADP-ribose) content.
RESULTS: A decrease (35%) in sensory nerve conduction velocity (SNCV) was seen in STZ-induced diabetic rats from day 10 post-STZ administration and persisted at days 25 and 39. At study completion (day 41), a decrease (32%) in intraepidermal nerve fiber density (IENFD) was found in hind-paw skin biopsies from STZ-rats. Reduced SNCV and IENFD were significantly ameliorated by both doses of Actovegin®. More-over, 600 mg/kg Actovegin® markedly decreased poly(ADP-ribose) polymerase (PARP) activity in sciatic nerves from STZ-diabetic rats as assessed by poly(ADP-ribose) content.
CONCLUSION: Actovegin® improved several para-meters of experimental diabetic neuropathy via mechanisms involving suppression of PARP activation, providing a rationale for treatment of this disease in humans.
SLV330, a cannabinoid CB1 receptor antagonist, ameliorates deficits in the T-maze, object recognition and Social Recognition Tasks in rodents
Cannabinoid CB(1) receptor (CB(1)R) signaling has been suggested to play an important role in the regulation of memory and cognition. In the present study, our aim was to investigate whether the CB(1)R antagonist SLV330 (doses ranging from 0.3 to 10mg/kg, given orally, p.o.) could ameliorate impairments in distinct aspects of cognition using different disruption models in both mice and rats. Effects of SLV330 were tested on working memory deficits in the T-maze Continuous Alternation Task (T-CAT) in mice; episodic memory deficits in the Object Recognition Task (ORT) and Social Recognition Task (SRT) in rats. The acetylcholinesterase inhibitor (AChEI) donepezil (Aricept, approved for symptomatic treatment of Alzheimer's disease) and nicotine were used as reference compounds. SLV330 markedly improved aging and scopolamine-induced memory deficits in the T-CAT in mice with a lowest effective dose (LED) of 1mg/kg p.o., while reversing the cognitive dysfunction induced by the N-methyl-D-aspartate (NMDA) antagonist dizocilpine (MK-801) only at the middle dose of 3mg/kg. In the ORT, we have found that combined administration of subthreshold doses of SLV330 (1mg/kg, p.o.) and the AChEI donepezil (0.1mg/kg, p.o.), that had no discernable effects on performance when given alone, enhanced memory performance in Wistar rats with deficits induced by the muscarinic antagonist scopolamine, suggestive of additive synergistic effects of SLV330 and donepezil on cognitive impairment. Finally, SLV330 was found to have cognition enhancing properties in a time delay paradigm in the SRT at a LED dose of 3mg/kg (p.o.). In conclusion, the CB(1)R antagonist SLV330 was found to clearly improve memory in several preclinical models for cognitive impairment.
Neuromuscular defects and breathing disorders in a new mouse model of spinal muscular atrophy
Spinal muscular atrophy (SMA) is caused by insufficient levels of the survival motor neuron (SMN) protein leading to muscle paralysis and respiratory failure. In mouse, introducing the human SMN2 gene partially rescues Smn(-)(/)(-) embryonic lethality. However current models were either too severe or nearly unaffected precluding convenient drug testing for SMA. We report here new SMN2;Smn(-/-) lines carrying one to four copies of the human SMN2 gene. Mice carrying three SMN2 copies exhibited an intermediate phenotype with delayed appearance of motor defects and developmental breathing disorders reminiscent of those found in severe SMA patients. Although normal at birth, at 7 days of age respiratory rate was decreased and apnea frequency was increased in SMA mice in parallel with the appearance of neuromuscular junction defects in the diaphragm. With median survival of 15 days and postnatal onset of neurodegeneration, these mice could be an important tool for evaluating new therapeutics. download
Animal Models of Collagen-Induced Arthritis
Collagen-induced arthritis in rats is associated with inflammatory polyarthritis, sharing clinical and pathological features with those of human rheumatoid arthritis (RA). Described in this unit is a protocol for consistently inducing arthritis in female Lewis rats by immunizing them with bovine type II collagen (CII) emulsified in complete Freund's adjuvant. This model is of value not only in defining the underlying pathogenesis of RA, but also as a tool for evaluating pharmacological strategies for treating this condition.
Specific Antinociceptive Activity of Cholest-4-en-3-one, Oxime (TRO19622) in Experimental Models of Painful Diabetic and Chemotherapy-Induced Neuropathy
Diabetes and cancer chemotherapies are often associated with painful neuropathy. The mechanisms underlying neuropathic pain remain poorly understood, and the current therapies have limited efficacy and are associated with dose-limiting side effects. We recently described the pharmacological characterization of cholest-4-en-3-one, oxime (TRO19622), a cholesterol-like compound, that significantly reduced axonal degeneration and accelerated recovery of motor nerve conduction in a model of peripheral neuropathy induced by crushing the sciatic nerve. These results triggered investigation of efficacy in other preclinical models of peripheral neuropathy. Here, we report evidence that daily oral administration of TRO19622, while similarly improving motor nerve conduction impaired in streptozotocin-induced diabetic rats, also reversed neuropathic pain behavior as early as the first administration. Further exploration of these acute antinociceptive effects demonstrated that TRO19622 was also able to reverse tactile allodynia in vincristine-treated rats, a model of chemotherapy-induced neuropathic pain. It is interesting to note that TRO19622 did not have analgesic activity in animal models of pain produced by formalin injection, noxious thermal or mechanical stimulation, or chronic constriction injury of the sciatic nerve, indicating that painful diabetic or chemotherapy-induced neuropathies share a common mechanism that is distinct from acute, inflammationdriven, or lesion-induced neuropathic pain. These results support the potential use of TRO19622 to treat painful diabetic and chemotherapy-induced neuropathies.
Interleukin-6 protects against paclitaxel, cisplatin and vincristine-induced neuropathies without impairing chemotherapeutic activity
PURPOSE: This study was conducted to investigate the potential neuroprotective effect of IL-6 on chemotherapy induced neuropathy (CIN). IL-6 was compared to four-methylcatechol (4-MC)-a known inducer of NGF secretion previously shown to exhibit neuroprotective effects in CIN models.
METHODS: Three CIN models were used; two in rats (cisplatin and vincristine) and one in mice (paclitaxel). IL-6 was delivered in four different doses in rats (0.3, 1, 3, 10 microg/kg, sc) every day from the first day of chemotherapeutic agent intoxication until the end of the study (day 37 for cisplatin protocol and day 30 for vincristine procedure). In mice, IL-6 was delivered at 10 microg/kg, sc either daily or three times a week from the first day of intoxication until the end of the study (day 19). Behavioral testings (hot plate and rotarod), nerve conduction studies (CMAP, SNCV, H-wave) and histo-morphometric analysis were done for all models. In addition, we tested whether IL-6 interfered with the tumor-reducing effects of the chemotherapeutic agents.
RESULTS: IL-6 treatment prevented the behavioral and electrophysiological abnormalities produced by vincristine, cisplatin and Taxol intoxication, and similarly prevented the pathological changes in peripheral nerves. The neuroprotective action of chronic IL-6 treatment was at least equal to that of 4-MC. In addition, IL-6 neither inhibited the antitumour activity of cisplatin, nor stimulated tumour growth. CONCLUSION: IL-6 at low doses (10 microg/kg) provided protection against the development of CIN without demonstrating interference with the anti tumoural activity of these anti-mitotic drugs.
Interleukin-6 attenuates the development of experimental diabetes-related neuropathy
Neuropathy is the most severe and the least understood complication of diabetes. We investigated the potential neuroprotective effect of IL-6 therapy in an experimental model of diabetic neuropathy. A single i.v. injection of streptozotocin (STZ, 55 mg/kg) was used to induce experimentaldiabetes in adult males. IL-6 (1, 10 or 30 microg/kg) was administrated either intraperitoneally on a daily basis or subcutaneously (s.c.) on a daily, on a three times or one time per week basis, starting at day 10 post-STZ. A decrease in sensory nerve conduction velocity (SNCV), indicative of neuropathy, is seen in STZ rats as early as day 10 post-STZ, a time at which blood glycaemia is already maximal. At later time points, this electrophysiological impairment became severe and clinically apparent by affecting tail flick latency. Motor dysfunction defined by a significant increase in compound muscle action potential (CMAP) latency was also recorded. At the completion of the study (day 40 post-STZ), histological examination revealed significant axonopathy and myelin loss, along with an increase in the proportion of fibers with abnormal appearance in sciatic nerves of STZ rats. These changes were not observed in non-diabetic rats and were significantly prevented by IL-6 treatment. The optimal dose appeared to be 10 microg/kg s.c. three injections per week, which showed a better effect in most of the parameters studied than 4-methylcatechol, a NGF-like neuroprotective compound. Once weekly and three times weekly administrations of IL-6 were as effective as daily treatment. Taken together, these results support the potential neuroprotective actions of IL-6. The fact that the half-life of IL-6 is only approximately 5 h while weekly dosing was neuroprotective strongly suggests activation by IL-6 of effector molecule(s) with longer duration of action. download
Functional maturation of nicotinic acetylcholine receptors as an indicator of murine muscular differentiation in a new nerve-muscle co-culture system
Under normal conditions in situ, muscle fibers and motoneurons, the main partners of motor units, are strongly dependent on each other. This interdependence hinders ex vivo studies of neuromuscular disorders where nervous or muscular components are considered separately. To allow in vitro access to complex nerve-muscle relationships, we developed a novel nerve-muscle co-culture system where mouse muscle innervation is assured by rat spinal cord explants. The degree of muscular maturation during co-culture was evaluated using the distribution of nicotinicacetylcholine receptors (AChRs) and their electrophysiological characteristics before and after innervation. In myotubes from non-innervated cultures, AChRs were diffusely distributed over the entire myotube surface. Their single-channel conductance (33.5+/-0.6 pS) and mean open time (8.1+/-0.7 ms) are characteristic of AChRs described in embryonic or denervated skeletal muscles. In innervated muscle fibers from co-cultures, AChRs appear as discrete aggregates and co-localize with synaptotagmin. In addition to the embryonic type currents, in innervated fibers AChR currents having high conductance (53.3+/-5.9 pS) and short mean open time (2.6+/-0.1 ms), characteristic of AChRs at mature neuromuscular junctions, were observed. Our data support the use of this new nerve-muscle co-culture system as a reliable model for the study of murine muscular differentiation and function.
Normal innervation and differentiation of X-linked myotubular myopathy muscle cells in a nerve-muscle coculture system
To study the pathogenesis of X-linked recessive myotubular myopathy (XLMTM), we used a nerve-muscle coculture system which allows the reconstitution of functional motor units in vitro after coupling of human skeletal muscle cells with embryonic rat spinal cord explants. We used three skeletal muscle cell lines derived from subjects with known mutations in the MTM1 gene (two from embryonic tissues, associated with mutations predicted to give a severe phenotype, and one from a neonate still alive at 3 years 6 months and exhibiting a mild phenotype). We compared these three XLMTM muscle cell cultures with control cultures giving special attention to behaviour of living cocultures (formation of the myofibres, contractile activity, survival), expression of muscular markers (desmin, dystrophin, alpha-actinin, troponin-T, myosin heavy chain isoforms), and nerve-muscle interactions (expression and aggregation of the nicotinic acetylcholine receptors). We were unable to reproduce any 'myotubular' phenotype since XLMTM muscle cells behaved like normal cells with regard to all the investigated parameters. Our results suggest that XLMTM muscle might be intrinsically normal and emphasize the possible involvement of the myotubularin-deficient motor neurons in the development of the disease. download
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- Preclinical CRO for CNS and PNS disorders
- In vivo and in vitro validated models
- In vivo and in vitro validated tests