Alternating-lever Cyclic-ratio Schedule Research Articles

MRZ-99030 – A novel modulator of Aβ aggregation: II – Reversal of Aβ oligomer-induced deficits in long-term potentiation (LTP) and cognitive performance in rats and mice

β-amyloid1-42 (Aβ1-42) is a major endogenous pathogen underlying the aetiology of Alzheimer's disease (AD). Recent evidence indicates that soluble Aβ oligomers, rather than plaques, are the major cause of synaptic dysfunction and neurodegeneration. Small molecules that suppress Aβ aggregation, reduce oligomer stability or promote off-pathway non-toxic oligomerization represent a promising alternative strategy for neuroprotection in AD. MRZ-99030 was recently identified as a dipeptide that modulates Aβ1-42 aggregation by triggering a non-amyloidogenic aggregation pathway, thereby reducing the amount of intermediate toxic soluble oligomeric Aβ species. The present study evaluated the relevance of these promising results with MRZ-99030 under pathophysiological conditions i.e. against the synaptotoxic effects of Aβ oligomers on hippocampal long term potentiation (LTP) and two different memory tasks. Aβ1-42 interferes with the glutamatergic system and with neuronal Ca2+ signalling and abolishes the induction of LTP. Here we demonstrate that MRZ-99030 (100–500 nM) at a 10:1 stoichiometric excess to Aβ clearly reversed the synaptotoxic effects of Aβ1-42 oligomers on CA1-LTP in murine hippocampal slices. Co-application of MRZ-99030 also prevented the two-fold increase in resting Ca2+ levels in pyramidal neuron dendrites and spines triggered by Aβ1-42 oligomers. In anaesthetized rats, pre-administration of MRZ-99030 (50 mg/kg s.c.) protected against deficits in hippocampal LTP following i.c.v. injection of oligomeric Aβ1-42. Furthermore, similar treatment significantly ameliorated cognitive deficits in an object recognition task and under an alternating lever cyclic ratio schedule after the i.c.v. application of Aβ1-42 and 7PA2 conditioned medium, respectively. Altogether, these results demonstrate the potential therapeutic benefit of MRZ-99030 in AD.

Effect of Intracerebroventricular Injection of TiO<SUB>2</SUB> Nanoparticles on Complex Behaviour in the Rat

There are no data available on the behavioural effects of centrally administered nanoparticles in freely moving intact mammals. Consequently, in the current study male Sprague-Dawley rats were trained to respond under an alternating-lever cyclic-ratio (ALCR) schedule of food reinforcement. Under this schedule, ascending and descending sequences of fixed-ratio (FR) lever press requirements for food reinforcement were presented over six cycles, with each discrete FR component completed on the alternate lever to the previous component. The final version of the schedule was comprised of an ascending followed by a descending sequence of the ratio values 2, 6, 12, 20, 30, 42 and 56, repeated over six cycles. When the rats were able to complete this version of the ALCR schedule in 40 min, each was implanted with a permanently indwelling ICV cannula aimed at the lateral ventricle of the brain, and allowed to recover for 7 days. On the first day of the experiment, all rats were injected with either titanium dioxide (TiO2, 9 nm, stabilised with gallic acid, 10 microl volume, 2 mg/ml) nanoparticles, or 10 microl saline (control). Two-hours after the ICV injections, the behaviour of all rats was measured using the ALCR schedule, and their behaviour was also measured (no ICV injection) for the next 7 days. Under the ALCR schedule, the number of lever-switching errors and incorrect lever perseverations significantly increased in the TiO2 group (p < 0.05). Other parameters of the ALCR schedule (RRRs and PRPs), which indicate the induction of malaise or general motor retardation, were not altered following ICV TiO2 injection. The findings of the current study indicate that central administration of TiO2 nanoparticles induced behavioural deterioration in freely moving intact animals, that the induced behavioural deterioration was a result of central rather than peripheral outcomes, and that this effect was chronic rather than acute.

Novel Anti-Inflammatory Compound SEN1176 Alleviates Behavioral Deficits Induced Following Bilateral Intrahippocampal Injection of Aggregated Amyloid-β1-42

Behavioral effects of a novel anti-inflammatory SEN1176 were investigated. This pyrrolo[3,2-e][1,2,4]triazolo[1,5-a]pyrimidine suppresses amyloid-β (Aβ)1-42-induced macrophage production of nitric oxide, TNF-α, IL-1β, and IL-6 in a dose-dependent fashion, an activity profile consistent with SEN1176 being a neuroinflammation inhibitor. Using male Sprague-Dawley rats, SEN1176 was examined relative to detrimental behavioral effects induced following bilateral intrahippocampal (IH) injections of aggregated Aβ1-42. The rats were trained to respond under an alternating-lever cyclic-ratio (ALCR) schedule of food reinforcement, enabling measurement of parameters of operant performance that reflect aspects of learning and memory. Under the ALCR schedule, orally administered SEN1176 at 5, 20, or 30 mg/kg was effective in reducing the behavioral deficit caused by bilateral IH aggregated Aβ1-42 injections in a dose-related manner over a 90-day treatment period. SEN1176 at 20 and 30 mg/kg significantly reduced lever switching errors and, at doses of 5, 10, and 30 mg/kg, significantly reduced incorrect lever perseverations, indicating a reduction of the behavioral deficit induced as a result of inflammation following IH Aβ1-42 injections. When treatment with SEN1176 was instigated 30 days after IH Aβ1-42 injections, it resulted in progressive protection, and withdrawal of SEN1176 treatment 60 days after IH Aβ1-42 injections revealed partial retention of the protective effect. SEN1176 also significantly reduced numbers of activated astrocytes adjacent to the aggregated Aβ1-42 injection sites. These results indicate the potential of SEN1176 for alleviating chronic neuroinflammatory processes related to brain Aβ deposition that affect learning and memory in Alzheimer's disease.

RS-0406 Arrests Amyloid-β Oligomer-Induced Behavioural Deterioration In Vivo

Clinically accessible compounds that arrest or reverse the effects of amyloid-β (Aβ) on progressively developing behavioural symptomatology and neuropathology in Alzheimer's disease (AD) have yet to become available. However, a viable strategy may be to target and neutralise soluble Aβ oligomers, which have been shown to mediate synaptic dysfunction and to produce cognitive deficits in the intact organism. Inhibiting the aggregation of Aβ is therapeutically attractive, as Aβ aggregation is a pathological event and pharmacological interventions targeting this are likely to have a non-toxic profile. A behavioural assay, the alternating-lever cyclic-ratio schedule, was used to assess the effect of Aβ oligomers and the non-peptide small molecule RS-0406 in male Sprague-Dawley rats. RS-0406 has been shown to inhibit Aβ1-42 fibrillogenesis and protect against Aβ1-42–induced cytotoxicity in primary hippocampal neurons. In the current study, RS-0406 ameliorated the adverse effects of secreted oligomers of human Aβ on behaviour and dose dependently reduced the behavioural effects of Aβ oligomers, with the highest dose, 10μM, maintaining behaviour approximately at control levels. This effect appeared to be central; peripheral confounds having been extensively investigated. This is the first published report on the effects of RS-0406 in vivo and indicates that RS-0406 has potential as a pharmacotherapeutic intervention for behavioural deficits seen in the early stages of AD, and possibly as an intervention in the development of AD neuropathology. Indeed, an analogue of RS-0406 that could be administered peripherally might be a realistic candidate for the clinical treatment of AD.

Bilateral intrahippocampal NAC 61–95 effects on behavior and moderation with l-NAME treatment

Residues 61–95 of the non-amyloid component (NAC 61–95) domain of α-synuclein are responsible for the aggregation and neurotoxicity of this protein. This study evaluated the effect of N (ω)-nitro- l-arginine methyl ester ( l-NAME), a nitric oxide synthase inhibitor, on the behavior of rats bilaterally injected into the CA3 area of the dorsal hippocampus with aggregated NAC 61–95. Twenty-four male Sprague–Dawley rats were trained to respond under an alternating-lever cyclic-ratio (ALCR) operant schedule. When responding was stable, 12 rats were injected bilaterally into the CA3 area of the hippocampus with aggregated NAC 61–95 (5 μl per side; concentration 10 −4 M), the remaining 12 rats were similarly injected with sterile water (SW), six of the NAC 61–95 injected rats and six of the SW injected rats were treated for 90 days post-injections with 0.05 mg/ml l-NAME in drinking water available ad libitum. Treatment with l-NAME alleviated NAC 61–95-induced behavioral deficits, indicated through the measurement of lever switching errors and incorrect lever perseverations under the ALCR schedule, and reduced the number of activated astrocytes proximal to the injection sites.

Congenital hypothyroidism impairs response alternation discrimination behavior

The behavior of six congenitally hypothyroid and six normal control rats was assessed under forced alternation fixed-ratio, alternating lever cyclic-ratio (ALCR) and progressive-ratio schedules of reinforcement. Hypothyroidism was produced by adding methimazole (MMI) to the drinking water of pregnant dams from embryonic day 16 to postnatal day 25. There were no differences in behavioral performance between MMI-treated and control animals under the fixed-ratio and progressive ratio schedules. There were also no differences in circulating triiodothyronine levels between groups at the end of the study. Under the ALCR schedule, when alternation of responding was forced during the first three cycles but both levers (choice) were presented during the last three cycles (correct lever active), the entire control group reached a competency criteria in nine sessions. In contrast, only two MMI-treated animals reached criteria after 17 sessions, and the remaining four MMI-treated animals did not reach criteria by 30 sessions of training. These results suggest that congenital hypothyroidism impairs learning when a discrimination between correct and incorrect operanda is made available.

Delayed behavioral effects following intrahippocampal injection of aggregated Aβ (1–42)

Beta amyloid protein (Aβ) is the major extracellular component of Alzheimer's disease (AD) plaques. In the current study, Aβ (1–42) was aggregated in vitro using a method which produces Aβ aggregates similar to those found in the AD brain. Twelve male Sprague–Dawley rats were trained in two-lever operant chambers under an alternating lever cyclic-ratio (ALCR) schedule. When performance was stable on the ALCR schedule, six subjects were injected (bilaterally into the CA3 area of the dorsal hippocampus) with 5.0 μl aggregated Aβ in suspension, and the remaining six subjects were injected with 5.0 μl sterile water. Behavioral testing resumed 5 days after surgery and continued for 90 days post-injection. Aggregated Aβ injection did not affect the number of lever switching errors made in a daily session but did affect the number of incorrect lever response perseverations. After approximately 30 days post-injection, aggregated Aβ injection detrimentally affected ability to track the changing parameters of the schedule, and decreased the efficiency by which subjects obtained reinforcers. From approximately day 50 post-injection onward, Aβ-injected subjects demonstrated significantly higher numbers of incorrect lever response perseverations than did sterile water-injected subjects. These effects appeared to be central rather than peripheral, as Aβ injection did not decrease running response rates under the ALCR schedule. The delayed onset of behavioral effects seen in this and other behavioral studies may be a result of a cascade of potentially harmful responses induced through glial activation following aggregated Aβ injection.