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Rationale Therapeutic agents for memory enhancement in psychiatric disorders, such as schizophrenia, are urgently needed. Objective The aim of this study is to characterize the preclinical profile of ITI-214, a potent inhibitor of phosphodiesterase 1 (PDE1). Methods ITI-214 was assayed for inhibition of PDE1 versus other PDE enzyme families using recombinant human PDE enzymes and for off-target binding to 70 substrates (General SEP II diversity panel; Caliper Life Sciences). Effects of ITI-214 (0.1–10 mg/kg, po) on memory performance were assayed in rats using the novel object recognition (NOR) paradigm, with drug given at specified time points prior to or following exposure to objects in an open field. ITI-214 was evaluated for potential drug-drug interaction with risperidone in rats using conditioned avoidance response (CAR) and pharmacokinetic assessments. Results ITI-214 inhibited PDE1A (Ki = 33 pmol) with >1000-fold selectivity for the nearest other PDE family (PDE4D) and displayed minimal off-target binding interactions in a 70-substrate selectivity profile. By using specific timing of oral ITI-214 administration, it was demonstrated in the NOR that ITI-214 is able to enhance acquisition, consolidation, and retrieval memory processes. All memory effects were in the absence of effects on exploratory behavior. ITI-214 did not disrupt the risperidone pharmacokinetic profile or effects in CAR. Conclusions ITI-214 improved the memory processes of acquisition, consolidation, and retrieval across a broad dose range (0.1–10 mg/kg, po) without disrupting the antipsychotic-like activity of a clinical antipsychotic medication, specifically risperidone. Clinical development of ITI-214 is currently in progress.

In previous studies, we have shown that phosphodiesterase type 5 inhibitors (PDE5-Is) can improve early consolidation of object memory. These conclusions were based on the timing of drug administration relative to the learning trial (i.e. before or after). However, there are very little pharmacological data available about the pharmacokinetic profile of orally administered PDE5-Is in the rat. Furthermore, there is still debate whether these effects are achieved via central or peripheral mechanisms and if acquisition processes are improved. In the current study, we tested the effects of the PDE5-I vardenafil in a cholinergic-deficit model and compared the effects after intracerebroventricular (ICV) versus oral (PO) administration. We found that PO vardenafil restored a scopolamine-induced memory impairment when dosed within 2 min after the learning trial while ICV vardenafil was able to restore memory when injected within 4 min after learning. Because the test trial was within 10 min after the learning trial, this suggests that these effects on object memory are related to acquisition processes that may still be ongoing in a time window after the learning trial. To further elucidate the extent of this acquisition window, we investigated the pharmacokinetic profile of vardenafil after PO administration where it was detected within 4 min post-dose. Taken together, our data suggest that PDE5 is involved in acquisition processes, which may linger for at least 4–6 min after learning. Further studies are needed to exclude that these effects could also be explained on basis of an effect on early consolidation processes. Additionally, the effectiveness of ICV-administered vardenafil provides further experimental evidence that PDE5-Is improve memory via a central mechanism.

Enhancement of central availability of the second messenger CAMP is a promising approach to improve cognitive function. Pharmacological inhibition of phosphodiesterase type 4 (PDE4), a group of cAMP hydrolyzing enzymes in the brain, has been shown to improve cognitive performances in rodents and monkeys. However, inhibition of PDE4 is generally associated with severe emetic side-effects. Roflumilast, an FDA-approved PDE4 inhibitor for treatment of chronic obstructive pulmonary disease (COPD), is yielding only mild emetic side effects. In the present study we investigate the potential of roflumilast as a cognition enhancer and to determine the potential coinciding emetic response in comparison to rolipram, a classic PDE4 inhibitor with pronounced emetic effects. Cognition enhancement was evaluated in mice and it was found that both roflumilast and rolipram enhanced memory in an object location task (0.03 mg/kg), whereas only roflumilast was effective in a spatial Y-maze (0.1 mg/kg). Emetic potential was measured using competition of PDE4 inhibition for alpha 2-adrenergic receptor antagonism in which recovery from xylazine/ketamine-mediated anesthesia is used as a surrogate marker. While rolipram displayed emetic properties at a dose 10 times the memory-enhancing dose, roflumilast only showed increased emetic-like properties at a dose 100 times the memory-enhancing dose. Moreover, combining sub-efficacious doses of the approved cognition enhancer donepezil and roflumilast, which did not improve memory when given alone, fully restored object recognition memory deficit in rats induced by the muscarinic receptor antagonist scopolamine. These findings suggest that roflumilast offers a more favorable window for treatment of cognitive deficits compared to rolipram. (C) 2016 Elsevier B.V. All rights reserved. Takeda (Deerfield, USA) funded the combination therapy study and participated in its design.

A promising target for memory improvement is phosphodiesterase type 5 (PDE5), which selectively hydrolyzes cyclic guanosine monophosphate (cGMP). In rodents, PDE5 inhibitors (PDE5-Is) have been shown to improve memory performance in many behavioral paradigms. However, it is questioned whether the positive effects in animal studies result from PDE5 inhibition in the central nervous system or the periphery. Therefore, we studied the effects of PDE5 inhibition on memory and determined whether compound penetration of the blood-brain barrier (BBB) is required for this activity. Two selective PDE5-Is, vardenafil and UK-343,664, were tested in the object recognition task (ORT) in both a MK-801- and scopolamine-induced memory deficit model, and a time-delay model without pharmacological intervention. Compounds were dosed 30 min before the learning trial of the task. To determine if the PDE5-Is crossed the BBB, their concentrations were determined in plasma and brain tissue collected 30 min after oral administration. Vardenafil improved object recognition memory in all three variants of the ORT. UK-343,664 was ineffective at either preventing MK-801-induced memory disruption or time-dependent memory decay. However, UK-343,664 attenuated the memory impairment of scopolamine. Vardenafil crossed the BBB whereas UK-343,664 did not. Further, co-administration of UK-343,664 and scopolamine did not alter the brain partitioning of either molecule. This suggests that the positive effect of UK-343,664 on scopolamine-induced memory decay might arise from peripheral PDE5 inhibition. The results herein suggest that there may be multiple mechanisms that mediate the efficacy of PDE5 inhibition to improve memory performance in tasks such as the ORT and that these involve PDE5 located both within and outside of the brain. To further elucidate the underlying mechanisms, the cellular and subcellular localization of PDE5 needs to be determined.

The object recognition task (ORT) allows assessing learning and memory processes in rodents. In this study, two areas in which knowledge about the ORT could be extended were addressed; i.e. generality to species and strains, and intervening variables including housing and estrous cycle. Regarding generality to species and strains, the ORT performance of golden hamsters was assessed. The hamsters showed sufficient exploration times, object recognition performance, and a retention-interval dependent decline similar to rats and mice. Subsequently, we tested three mouse strains which have not been described before in the ORT; i.e. OF1, NMRI, and SJL mice. OF1 and NMRI strains performed equally well, whereas the SJL strain showed low exploration times and no memory retention. Therefore, the SJL strain is unsuited for ORT experiments using a 1 h retention interval and a fixed (3 min) trial duration. Furthermore, the sensitivity to a pharmacological memory deficit model (scopolamine) was tested in three rat strains. Each strain showed a dose dependent relationship, but the least effective dose of scopolamine differed among the three strains, the effect being greater in the order of Wistar, Long-Evans, Hooded Lister rats. Finally, to investigate potential intervening variables in the ORT, the effects of housing conditions and estrous cycle were investigated with rats. Single housing resulted in absolute higher performance than social housing. Furthermore, females in pro-estrus/estrus showed better performance compared to females in metestrus/di-estrus. Taken together, object recognition appears to be a common ability of rodent species, but different strains have different memory capacities and sensitivities to scopolamine, individual housing leads to higher performance, and performance of females is dependent on the estrous cycle phase. Thus, rodent species, strain, housing, and estrous cycle should be taken into consideration in ORT studies. (C) 2012 Elsevier B.V. All rights reserved.

The 5-hydroxytryptamine(6) (5-HT(6)) receptor 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 novel, selective 5-HT(6) antagonists compound (CMP) X and CMP Y and the reference 5-HT(6) antagonist GSK-742457 could ameliorate impairments in episodic memory in 3-months-old male Wistar rats. The acetylcholinesterase inhibitor (AChEI) donepezil (Aricept®, approved for symptomatic treatment of Alzheimer's disease, AD) was used as a positive reference compound. First, effects of the 5-HT(6) antagonists CMP X, CMP Y and GSK-742457 were investigated on object recognition task (ORT) performance in rats treated with the muscarinic antagonist scopolamine (0.1mg/kg, administered intraperitoneally, i.p., 30 min before trial 1). Second, effects of the combination of suboptimal doses of 5-HT(6) antagonists CMP X and CMP Y with the AChEI donepezil were studied, to determine whether the 5-HT(6) antagonists show additive synergism with donepezil in the ORT. Finally, effects of CMP Y, GSK-742457 and donepezil were investigated on object location task (OLT) performance in rats treated with scopolamine. Donepezil (1mg/kg, oral administration, p.o.), GSK-742457 (3mg/kg, i.p.), CMP X (3mg/kg, i.p.) and CMP Y (30 mg/kg, p.o.), all ameliorated the scopolamine-induced deficits in object recognition. In the ORT, we have found that combined administration of subthreshold doses of CMP X (1mg/kg, i.p.) and CMP Y (10mg/kg, p.o.) with the AChEI donepezil (0.1mg/kg, p.o.), enhanced memory performance in Wistar rats with deficits induced by scopolamine. Donepezil (0.1mg/kg, p.o.) alone had no discernable effects on performance. This suggests additive synergistic effects of the 5-HT(6) antagonists (CMP X and CMP Y) with donepezil on cognitive impairment. Finally, donepezil (1mg/kg, p.o.), GSK-742457 (10mg/kg, p.o.) and CMP Y (30 mg/kg, p.o.) also reduced scopolamine-induced deficits in the OLT. In conclusion, the 5-HT(6) antagonists were found to clearly improve episodic memory deficits induced by scopolamine. In addition, co-administration of the 5-HT(6) receptor antagonists CMP X and CMP Y with the AChEI donepezil to cognitively impaired rats also resulted in potentially additive enhancing effects on cognition. This suggests that these compounds could have potential as monotherapy, but also as adjunctive therapy in patients with AD treated with common treatments such as donepezil.

Cannabinoid CB1 receptor (CB1R) 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 CB1R antagonist SLV330 (doses ranging from 0.3 to 10 mg/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 1 mg/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 3 mg/kg. In the ORT, we have found that combined administration of subthreshold doses of SLV330 (1 mg/kg, p.o.) and the AChEI donepezil (0.1 mg/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 3 mg/kg (p.o.). In conclusion, the CB1R antagonist SLV330 was found to clearly improve memory in several preclinical models for cognitive impairment.

The second messengers cGMP and cAMP have a vital role in synaptic plasticity and memory processes. As such, phosphodiesterases inhibitors (PDE-Is), which prevent the breakdown of these cyclic nucleotides, represent a potential treatment strategy in memory decline. Recently it has been demonstrated that cGMP and cAMP signaling act in sequence during memory consolidation, with early cGMP signaling requiring subsequent cAMP signaling. Here, we sought to confirm this relationship, and to evaluate its therapeutic implications. Combining sub-efficacious doses of the cGMP-specific PDE type 5 inhibitor vardenafil (0.1 mg/kg) and cAMP-specific PDE type 4 inhibitor rolipram (0.01 mg/kg) during the early and late memory consolidation phase, respectively, led to improved memory performance in a 24 h interval object recognition task. Similarly, such a sub-efficacious combination treatment enhanced the transition of early-phase long-term potentiation (LTP) to late-phase LTP in hippocampal slices. In addition, both object memory and LTP were improved after administration of two sub-efficacious doses of the dual substrate PDE type 2 inhibitor BAY60 7550 (0.3 mg/kg) at the early and late consolidation phase, respectively. Taken together, combinations of sub-efficacious doses of cAMP- and cGMP-specific PDE-Is have an additive effect on long-term synaptic plasticity and memory formation and might prove a superior alternative to single PDE-I treatment.

Drug effects are usually evaluated in animals housed under maximally standardized conditions. However, it is assumed that an enriched environment (EE) more closely resembles human conditions as compared to maximally standardized laboratory conditions. In the present study, we examined the acute cognition enhancing effects of vardenafil, a PDE5 inhibitor, which stimulates protein kinase G/CREB signaling in cells, in three different groups of male Wistar rats tested in an object recognition task (ORT). Rats were either housed solitarily (SOL) or socially (SOC) under standard conditions, or socially in an EE. Although EE animals remembered object information longer in the vehicle condition, vardenafil only improved object memory in SOL and SOC animals. While EE animals had a heavier dorsal hippocampus, we found no differences between experimental groups in total cell numbers in the dentate gyrus, CA2-3 or CA1. Neither were there any differences in markers for pre- and postsynaptic density. No changes in PDE5 mRNA- and protein expression levels were observed. Basal pCREB levels were increased in EE rats only, whereas beta-catenin was not affected, suggesting specific activation of the MAP kinase signaling pathway and not the AKT pathway. A possible explanation for the inefficacy of vardenafil could be that CREB signaling is already optimally stimulated in the hippocampus of EE rats. Since previous data has shown that acute PDE5 inhibition does not improve memory performance in humans, the use of EE animals could be considered as a more valid model for testing cognition enhancing drugs.

We examined the temporal profile of pharmacologically enhanced episodic memory, using the object recognition task. Male Wistar rats were tested at different retention intervals ranging from 1 h to 24 h. The object discrimination performance of all groups (untreated, placebo, drug treatment) gradually decreased up to an interval (8 h). Interestingly, only after this 8 h interval the memory improving effects of vardenafil and rolipram started to emerge. This time-dependent memory performance shows similarities with the Kamin effect. The delayed manifestation of drug-enhanced memory suggests that two separate memory mechanisms are at play, a quick transient form of memory and a more stable memory form that requires several hours to develop. It is important to take this into account when testing treatments intended for long-term memory enhancement.

Objective To further explore the implication of the serotonin (5-HT) system in the improvement of rat short-term object recognition after administration of the type 2 phosphodiesterase inhibitor (PDE-I) BAY 60-7550 and the type 5 PDE-I vardenafil, the effect of PDE2 and PDE5 inhibition upon central amino acid levels, 5-HT, and related parameters were measured after applying acute tryptophan depletion (ATD). Method Wistar rats were orally administered saline or a protein–carbohydrate mixture with or without tryptophan (TRP). TRP-depleted animals additionally received an oral vehicle injection or the PDE inhibitors BAY 60–7550 or vardenafil at a dose known to improve object memory performance. Results Although ATD significantly decreased TRP levels in the hippocampus 2 h after administration, 5-HT levels appeared only moderately affected, without any changes observed in the amount of 5-HIAA or 5-HT turnover rate. Moreover, no effects of PDE inhibition upon 5-HT or related parameters were observed. Conclusion Changes in 5-HT neurotransmitter activity might be excluded as a potential underlying mechanism of the previously reported ability of PDE inhibitors to improve short-term object memory in rats. It is suggested that a decrease in cerebral blood flow potentially underlies ATD-induced object memory deficits, most likely due to decrease in NO synthesis.