M1/M4 receptors as potential therapeutic treatments for schizophrenia: A comprehensive study.

Muscarinic acetylcholine receptors (mAChRs) play a significant role in the pathophysiology of schizophrenia. Although activating mAChRs holds potential in addressing the full range of schizophrenia symptoms, clinical application of many non-selective mAChR agonists in cognitive deficits, positive and negative symptoms is hindered by peripheral side effects (gastrointestinal disturbances and cardiovascular effects) and dosage restrictions. Ligands binding to the allosteric sites of mAChRs, particularly the M1 and M4 subtypes, demonstrate activity in improving cognitive function and amelioration of positive and negative symptoms associated with schizophrenia, enhancing our understanding of schizophrenia. The article aims to critically examine current design concepts and clinical advancements in synthesizing and designing small molecules targeting M1/M4, providing theoretical insights and empirical support for future research in this field.

Balanced modulation of neuromuscular synaptic transmission via M1 and M2 muscarinic receptors during inhibition of cholinesterases.

Organophosphorus (OP) compounds that inhibit acetylcholinesterase are a common cause of poisoning worldwide, resulting in several hundred thousand deaths each year. The pathways activated during OP compound poisoning via overstimulation of muscarinic acetylcholine receptors (mAChRs) play a decisive role in toxidrome. The antidotal therapy includes atropine, which is a nonspecific blocker of all mAChR subtypes. Atropine is efficient for mitigating depression in respiratory control centers but does not benefit patients with OP-induced skeletal muscle weakness. By using an ex vivo model of OP-induced muscle weakness, we studied the effects of the M1/M4 mAChR antagonist pirenzepine and the M2/M4 mAChR antagonist methoctramine on the force of mouse diaphragm muscle contraction. It was shown that weakness caused by the application of paraoxon can be significantly prevented by methoctramine (1 µM). However, neither pirenzepine (0.1 µM) nor atropine (1 µM) was able to prevent muscle weakness. Moreover, the application of pirenzepine significantly reduced the positive effect of methoctramine. Thus, balanced modulation of neuromuscular synaptic transmission via M1 and M2 mAChRs contributes to paraoxon-induced muscle weakness. It was shown that methoctramine (10 µmol/kg, i.p.) and atropine (50 µmol/kg, i.p.) were equieffective toward increasing the survival of mice poisoned with a 2xLD50 dose of paraoxon.

Therapeutic potential of TAK-071, a muscarinic M1 receptor positive allosteric modulator with low cooperativity, for the treatment of cognitive deficits and negative symptoms associated with schizophrenia.

The selective activation of the muscarinic M1 receptor (M1R) may be a promising approach for treating cognitive impairment associated with cholinergic dysfunction. We previously reported that low cooperativity (α-value) is associated with a favorable cholinergic side effect profile of M1R positive allosteric modulators (M1 PAMs), as well as being a crucial factor for the cognitive improvement observed after combining M1 PAMs with donepezil, in rodents. In this study, we preclinically characterized TAK-071, a novel M1 PAM with low cooperativity (α-value = 199), as a new therapy for schizophrenia. We tested TAK-071 in the offspring of polyriboinosinic-polyribocytidylic acid-treated dams, which is a maternal immune activation model of schizophrenia. TAK-071 improved sociability deficits and working memory in this model. In a genetic mouse model of schizophrenia, miR-137 transgenic (Tg) mice, TAK-071 improved deficits in working memory, recognition memory, sociability, and sensorimotor gating. Patients with schizophrenia usually take several antipsychotics to treat positive symptoms. Thus, we also investigated the combined effects of TAK-071 with currently prescribed antipsychotics. Among the 10 antipsychotics tested, only olanzapine and quetiapine showed M1R antagonistic effects, which were counteracted by TAK-071 at possible effective concentrations for cognitive improvement in vitro. Moreover, haloperidol did not affect the ability of TAK-071 to improve working memory in miR-137 Tg mice, suggesting a low risk of losing efficacy when combined with dopamine D2 receptor antagonists. In conclusion, TAK-071 can exert beneficial effects on social behavior and cognitive function and could be a new therapy for schizophrenia.

Muscarinic M1, but not M4, receptor antagonism impairs divided attention in male rats.

Divided attention may be more important than ever to comprehend, given ubiquitous distractors in modern living. In humans, concern has been expressed about the negative impact of distraction in education, the home, and the workplace. While acetylcholine supports divided attention, in part via muscarinic receptors, little is known about the specific muscarinic subtypes that may contribute. We designed a novel, high-response rate test of auditory sustained attention, in which rats complete variable-ratio runs on one of two levers, rather than emitting a single response. By doing this, we can present a secondary visual distractor task during some trials, for which a correct nosepoke response is reinforced with a more palatable food pellet. The nonspecific muscarinic antagonist scopolamine impaired performance, and slowed and reduced lever press activity. We then explored antagonists that preferentially block the M1 and M4 subtypes, because these receptors are potential therapeutic targets for cognitive enhancers. Telenzepine, an M1-preferring antagonist, impaired divided attention performance, but not performance of the attention task without distraction. Telenzepine also had fewer nonspecific effects than scopolamine. In contrast, the M4-preferring antagonist tropicamide had no effects. Analysis of overall behavior also indicated that accuracy in the main attention task decreased as a function of engagement with the distractor task. These results implicate the M1 receptor in divided attention.

Effects of biperiden (cholinergic muscarinic m1/m4 receptor antagonist) on ethanol conditioned place preference in mice.

BACKGROUND: Previous studies suggest that muscarinic cholinergic receptors might act upon the dopamine release in the mesolimbic system and alter drug-reinforcing values related to drug craving. AIMS: We examined the effects of systemic biperiden administration, a muscarinic cholinergic (M1/M4) receptor antagonist, on ethanol (dose of 2 g/Kg) conditioned place preference (CPP), neuronal activation, dopamine and its metabolites levels in the nucleus accumbens. METHODS: Thirty minutes before the ethanol-induced CPP test, mice received saline or biperiden at doses of 1.0, 5.0, or 10.0 mg/kg. The time spent in each compartment was recorded for 15 min. After the CPP protocol, animals were euthanized, and we investigated the activation of the nucleus accumbens by immunohistochemistry for Fos. We also quantified dopamine, homovanillic acid (HVA), and dihydroxyphenylacetic acid (DOPAC) levels in the nucleus accumbens by high-performance liquid chromatography (HPLC). Additionally, the rotarod was employed to evaluate the effects of biperiden on motor coordination. RESULTS: Biperiden at different doses (1.0, 5.0, and 10.0 mg/kg) blocked the expression of ethanol-induced CPP. These biperiden doses increased the number of Fos-positive cells and the dopamine turnover in the nucleus accumbens. None of the doses affected the motor coordination evaluated by the rotarod. CONCLUSIONS: Our results show that biperiden can modulate the effect of alcohol reward, and its mechanism of action may involve a change in dopamine and cholinergic mesolimbic neurotransmission.

Striatal and nigral muscarinic type 1 and type 4 receptors modulate levodopa-induced dyskinesia and striato-nigral pathway activation in 6-hydroxydopamine hemilesioned rats.

Acetylcholine muscarinic receptors (mAChRs) contribute to both the facilitation and inhibition of levodopa-induced dyskinesia operated by striatal cholinergic interneurons, although the receptor subtypes involved remain elusive. Cholinergic afferents from the midbrain also innervate the substantia nigra reticulata, although the role of nigral mAChRs in levodopa-induced dyskinesia is unknown. Here, we investigate whether striatal and nigral M1 and/or M4 mAChRs modulate dyskinesia and the underlying striato-nigral GABAergic pathway activation in 6-hydroxydopamine hemilesioned rats. Reverse microdialysis allowed to deliver the mAChR antagonists telenzepine (M1 subtype preferring), PD-102807 and tropicamide (M4 subtype preferring), as well as the selective M4 mAChR positive allosteric modulator VU0152100 in striatum or substantia nigra, while levodopa was administered systemically. Dyskinetic movements were monitored along with nigral GABA (and glutamate) and striatal glutamate dialysate levels, taken as neurochemical correlates of striato-nigral pathway and cortico-basal ganglia-thalamo-cortical loop activation. We observed that intrastriatal telenzepine, PD-102807 and tropicamide alleviated dyskinesia and inhibited nigral GABA and striatal glutamate release. This was partially replicated by intrastriatal VU0152100. The M2 subtype preferring antagonist AFDX-116, used to elevate striatal acetylcholine levels, blocked the behavioral and neurochemical effects of PD-102807. Intranigral VU0152100 prevented levodopa-induced dyskinesia and its neurochemical correlates whereas PD-102807 was ineffective. These results suggest that striatal, likely postsynaptic, M1 mAChRs facilitate dyskinesia and striato-nigral pathway activation in vivo. Conversely, striatal M4 mAChRs can both facilitate and inhibit dyskinesia, possibly depending on their localization. Potentiation of striatal and nigral M4 mAChR transmission leads to powerful multilevel inhibition of striato-nigral pathway and attenuation of dyskinesia.

Blockade of the M1 muscarinic acetylcholine receptors impairs eyeblink serial feature-positive discrimination learning in mice.

The serial feature-positive discrimination task requires the subjects to respond differentially to the identical stimulus depending on the temporal context given by a preceding cue stimulus. In the present study, we examined the involvement of the M1 muscarinic acetylcholine receptors using a selective M1 antagonist VU0255035 in the serial feature-positive discrimination task of eyeblink conditioning in mice. In this task, mice received a 2-s light stimulus as the conditional cue 5 or 6 s before the presentation of a 350-ms tone conditioned stimulus (CS) paired with a 100-ms peri-orbital electrical shock (cued trials), while they did not receive the cue before the presentation of the CS alone (non-cued trials). Each day mice randomly received 30 cued and 30 non-cued trials. We found that VU0255035 impaired acquisition of the conditional discrimination as well as the overall acquisition of the conditioned response (CR) and diminished the difference in onset latency of the CR between the cued and non-cued trials. VU0255035 administration to the control mice after sufficient learning did not impair the pre-acquired conditional discrimination or the CR expression itself. These effects of VU0255035 were almost similar to those with the scopolamine in our previous study, suggesting that among the several types of muscarinic acetylcholine receptors, the M1 receptors may play an important role in the acquisition of the conditional discrimination memory but not in mediating the discrimination itself after the memory had formed in the eyeblink serial feature-positive discrimination learning.

Structure and selectivity engineering of the M1 muscarinic receptor toxin complex.

Muscarinic toxins (MTs) are natural toxins produced by mamba snakes that primarily bind to muscarinic acetylcholine receptors (MAChRs) and modulate their function. Despite their similar primary and tertiary structures, MTs show distinct binding selectivity toward different MAChRs. The molecular details of how MTs distinguish MAChRs are not well understood. Here, we present the crystal structure of M1AChR in complex with MT7, a subtype-selective anti-M1AChR snake venom toxin. The structure reveals the molecular basis of the extreme subtype specificity of MT7 for M1AChR and the mechanism by which it regulates receptor function. Through in vitro engineering of MT7 finger regions that was guided by the structure, we have converted the selectivity from M1AChR toward M2AChR, suggesting that the three-finger fold is a promising scaffold for developing G protein-coupled receptor modulators.

Activation of cortical M1 muscarinic receptors and related intracellular signaling is necessary for reactivation-induced object memory updating.

Reactivated long-term memories can become labile and sensitive to modification. Memories in this destabilized state can be weakened or strengthened, but there is limited research characterizing the mechanisms underlying retrieval-induced qualitative updates (i.e., information integration). We have previously implicated cholinergic transmission in object memory destabilization. Here we present a novel rodent paradigm developed to assess the role of this cholinergic mechanism in qualitative object memory updating. The post-reactivation object memory modification (PROMM) task exposes rats to contextual information following object memory reactivation. Subsequent object exploratory performance suggests that the contextual information is integrated with the original memory in a reactivation- and time-dependent manner. This effect is blocked by interference with M1 muscarinic receptors and several downstream signals in perirhinal cortex. These findings therefore demonstrate a hitherto unacknowledged cognitive function for acetylcholine with important implications for understanding the dynamic nature of long-term memory storage in the normal and aging brain.

T-495, a novel low cooperative M1 receptor positive allosteric modulator, improves memory deficits associated with cholinergic dysfunction and is characterized by low gastrointestinal side effect risk.

M1 muscarinic acetylcholine receptor (M1 R) activation can be a new therapeutic approach for the treatment of cognitive deficits associated with cholinergic hypofunction. However, M1 R activation causes gastrointestinal (GI) side effects in animals. We previously found that an M1 R positive allosteric modulator (PAM) with lower cooperativity (α-value) has a limited impact on ileum contraction and can produce a wider margin between cognitive improvement and GI side effects. In fact, TAK-071, a novel M1 R PAM with low cooperativity (α-value of 199), improved scopolamine-induced cognitive deficits with a wider margin against GI side effects than a high cooperative M1 R PAM, T-662 (α-value of 1786), in rats. Here, we describe the pharmacological characteristics of a novel low cooperative M1 R PAM T-495 (α-value of 170), using the clinically tested higher cooperative M1 R PAM MK-7622 (α-value of 511) as a control. In rats, T-495 caused diarrhea at a 100-fold higher dose than that required for the improvement of scopolamine-induced memory deficits. Contrastingly, MK-7622 showed memory improvement and induction of diarrhea at an equal dose. Combination of T-495, but not of MK-7622, and donepezil at each sub-effective dose improved scopolamine-induced memory deficits. Additionally, in mice with reduced acetylcholine levels in the forebrain via overexpression of A53T α-synuclein (ie, a mouse model of dementia with Lewy bodies and Parkinson's disease with dementia), T-495, like donepezil, reversed the memory deficits in the contextual fear conditioning test and Y-maze task. Thus, low cooperative M1 R PAMs are promising agents for the treatment of memory deficits associated with cholinergic dysfunction.

Central muscarinic receptor subtypes (M1 and M3) involved in carbacol-induced hypophagia in neonatal broiler chicken.

Aim: Food intake regulated by a complex of physiologic mechanisms in the nervous system. Muscarinergic system has an important role in the central regulation of appetite in mammals, but there is no information for Muscarinic receptors in avian. The purpose of this study was to examine the effects of intracerebroventricular injection of carbachol (cholinergic agonist), Telenzepine (M1 receptor antagonist), AF-DX116 (M2 receptor antagonist), 4-DAMP (M3 receptor antagonist), and PD102807 (M4 receptor antagonist) on feeding behavior in 3-h food-deprived (FD3) neonatal broiler chicken.Materials and Methods: In experiment 1, chicken intracerebroventricular injected with carbachol (125, 250, and 500 nmol). In experiment 2, birds intracerebroventricular injected with telenzepine (125, 250, and 500 nmol). In experiments 3-5, birds intracerebroventricular injected with AF-DX 116 (125, 250, and 500 nmol), 4-DAMP (125, 250, and 500 nmol), and PD102807 (125, 250, and 500 nmol), respectively. In experiment 6, broilers intracerebroventricular injected with carbacol (500 nmol), co-injection of telenzepine (125 nmol)+carbacol (500 nmol), and 4-DAMP (125 nmol)+carbacol (500 nmol). In experiment 7, injection procedure was carbacol (500 nmol), co-injection of AF-DX116 (125 nmol)+carbacol (500 nmol), and PD102807 (125 nmol)+carbacol (500 nmol). Then, food intake measured until 120 min after injection.Results: According to the data, carbachol (250 and 500 nmol) significantly decreased food intake in comparison with control group (P < 0.05). Intracerebroventricular injection of telenzepine (250 and 500 nmol) and 4-DAMP (250 and 500 nmol) significantly increased food intake (P < 0.05). In addition, carbacol-induced hypophagia was significantly attenuated by co-injection of telenzepine + carbacol (P < 0.05). Also, co-injection of 4-DAMP + carbacol decreased the effect of carbacol on food intake (P < 0.05). However, AF-DX116 and PD102807 had no effect on hypophagia induced by carbacol (P > 0.05).Conclusion: These results suggest, hypophagic effect of muscarinergic system is mediated via M1 and M3 receptors in neonatal chicken.

M1 muscarinic receptor is a key target of neuroprotection, neuroregeneration and memory recovery by i-Extract from Withania somnifera.

Memory loss is one of the most tragic symptoms of Alzheimer's disease. Our laboratory has recently demonstrated that 'i-Extract' of Ashwagandha (Withania somnifera) restores memory loss in scopolamine (SC)-induced mice. The prime target of i-Extract is obscure. We hypothesize that i-Extract may primarily target muscarinic subtype acetylcholine receptors that regulate memory processes. The present study elucidates key target(s) of i-Extract via cellular, biochemical, and molecular techniques in a relevant amnesia mouse model and primary hippocampal neuronal cultures. Wild type Swiss albino mice were fed i-Extract, and hippocampal cells from naïve mice were treated with i-Extract, followed by muscarinic antagonist (dicyclomine) and agonist (pilocarpine) treatments. We measured dendritic formation and growth by immunocytochemistry, kallikrein 8 (KLK8) mRNA by reverse transcription polymerase chain reaction (RT-PCR), and levels of KLK8 and microtubule-associated protein 2, c isoform (MAP2c) proteins by western blotting. We performed muscarinic receptor radioligand binding. i-Extract stimulated an increase in dendrite growth markers, KLK8 and MAP2. Scopolamine-mediated reduction was significantly reversed by i-Extract in mouse cerebral cortex and hippocampus. Our study identified muscarinic receptor as a key target of i-Extract, providing mechanistic evidence for its clinical application in neurodegenerative cognitive disorders.

A Selective M1 and M3 Receptor Antagonist, Penehyclidine Hydrochloride, Exerts Antidepressant-Like Effect in Mice.

Recent studies indicate that anti-muscarinic receptor is a prospective strategy to treat depression. Although non-selective antagonist of muscarinic receptor scopolamine exhibits rapid and robust antidepressant-like effect, it still has various side effects including abuse risk. Penehyclidine hydrochloride (PHC) is a novel clinical anti-cholinergic drug derived from scopolamine in China, which selectively blocks M1 and M3 muscarinic receptor. Therefore, the objective of this study was to evaluate whether PHC would manifest antidepressant-like effects. Forced swim test (FST), tail suspension test (TST) and chronic unpredictable mild stress (CUMS) model of depression were explored to assess the antidepressant-like effect. Western blotting was further performed to detect the effects of PHC on the brain-derived neurotrophic factor (BDNF) signal cascade. Immunofluorescence was used to observe the activation of astrocyte. Moreover, different pharmacological inhibitors were applied to clarify the antidepressant-like mechanism. The results of the present experiments revealed that PHC decreased the immobility time of FST and TST in mice. In the CUMS model, PHC rapidly ameliorated anhedonia-like behavior (within 4 days), accompanying with the enhanced expression of BDNF and phosphorylation of extracellular signal-related kinase 1/2 (ERK1/2) in the hippocampus. In addition, blockade of the BDNF release by verapamil and activation of its Trk B receptor by K252a, rather than inhibition of opioid system by naloxone or sigma receptor by BD1047, abolished the antidepressant-like effects of PHC in mice. The findings suggest that PHC, an anti-muscarinic drug in clinical use, elicits rapid onset antidepressant-like effect, shedding light on the development of new antidepressants.

Muscarinic M1 and M2 receptor subtypes play opposite roles in LPS-induced septic shock.

BACKGROUND: To compare pharmacologic effects of pirenzepine and AF-DX116, a selective competitive antagonist for M1 and M2 subtype muscarinic cholinergic receptors (mAChRs), respectively, with atropine, a non-selective competitive antagonist for mAChRs, on Lipopolysaccharide (LPS). METHODS: Male C57BL/6 mice were used to establish models of LPS-induced experimental endotoxemia. Mice were intraperitoneally injected 10 min prior to LPS injection with control (saline), atropine, pirenzepine and AF-DX116, respectively. Overall survival time was estimated using Kaplan-Meier plots. Inflammatory cytokine tumor necrosis factor-α (TNF-α) was monitored at various intervals after LPS injection and individual reagent administration. Pathological alternations in lungs and liver were analyzed. RESULTS: Pirenzepine and atropine pretreatment improved survival rate of LPS-induced septic shock; in contrast, AF-DX116 accelerated death from sepsis. Moreover, TNF-α plasma level was decreased in response to pirenzepine or atropine, whereas increased in response to AF-DX116. Pirenzepine and atropine relieved whereas AF-DX116 accelerated LPS-induced pulmonary and hepatic injury. Pirenzepine reduced proportion of M1 subtype of macrophages, while AF-DX116 promoted polarization of macrophages to M1 subtype. Pirenzepine pretreatment reduced while AF-DX116 enhanced expression of SOCS3 at mRNA level. CONCLUSIONS: The administration of pirenzepine and atropine may have beneficial effects on septic shock.

Role of Kv7.2/Kv7.3 and M1 muscarinic receptors in the regulation of neuronal excitability in hiPSC-derived neurons.

The Kv7 family of voltage-dependent non-inactivating potassium channels is composed of five members, of which four are expressed in the CNS. Kv7.2, 7.3 and 7.5 are responsible for the M-current, which plays a critical role in the regulation of neuronal excitability. Stimulation of M1 muscarinic acetylcholine receptor, M1 receptor, increases neuronal excitability by suppressing the M-current generated by the Kv7 channel family. The M-current modulation via M1 receptor is well-described in in vitro assays using cell lines and in native rodent tissue. However, this mechanism was not yet reported in human induced pluripotent stem cells (hiPSC) derived neurons. In the present study, we investigated the effects of both agonists and antagonists of Kv7.2/7.3 channel and M1 receptor in hiPSC derived neurons and in primary rat cortical neuronal cells. The role of M1 receptors in the modulation of neuronal excitability could be demonstrated in both rat primary and hiPSC neurons. The M1 receptors agonist, xanomeline, increased neuronal excitability in both rat cortical and the hiPSC neuronal cells. Furthermore, M1 receptor agonist-induced neuronal excitability in vitro was reduced by an agonist of Kv7.2/7.3 in both neuronal cells. These results show that hiPSC derived neurons recreate the modulation of the M-current by the muscarinic receptor in hiPSC neurons similarly to rat native neurons. Thus, hiPSC neurons could be a useful human-based cell assay for characterization of drugs that affect neuronal excitability and/or induce seizure activity by modulation of M1 receptors or inhibition of Kv7 channels.

Sialorrhea Successfully Treated by the Combined Use of Selective M1 and M3 Muscarinic Acetylcholine Receptor Antagonists.

Sialorrhea is often treated with anticholinergic agents, but they can have undesirable side effects such as drowsiness, sedation, and constipation. Effective medication that acts selectively on the salivary glands is needed. We report the case of a patient with sialorrhea who was successfully treated by the combined use of pirenzepine and solifenacin (M1 and M3 muscarinic receptor antagonists, respectively). The patient was a 51-year-old man with mean unstimulated and stimulated salivary flow rates per 10 min of 6.1 mL and 41.7 mL, respectively (both were measured three times). 99mTcO4- salivary gland scintigraphy revealed characteristic spontaneous saliva secretion without stimulation. He was treated with Scopolia extract, escitalopram, solifenacin succinate, and the combined administration of solifenacin succinate and pirenzepine. A statistically significant decrease was observed from the pre-medication unstimulated and stimulated salivary flow rates only following the combined administration of solifenacin and pirenzepine. The major muscarinic receptor subtype expressed in the salivary glands is M3; however, M1 is also present. A study using knockout mice demonstrated that the presence of either M1 or M3 receptors was sufficient for salivation. Thus, the combined use of selective M1 and M3 antagonists could provide a good treatment option for sialorrhea.

Effects of Muscarinic Acetylcholine m1 and m4 Receptor Blockade on Dyskinesia in the Hemi-Parkinsonian Rat.

Standard treatment for Parkinson's disease (PD) is L-DOPA, but with chronic administration the majority of patients develop L-DOPA-induced dyskinesia (LID). Emerging evidence implicates the cholinergic system in PD and LID. Muscarinic acetylcholine receptors (mAChR) are known to modulate movement and of late have been implicated as possible targets for LID. Therefore the current study investigated the role of M1 and M4 mAChRs in LID, on motor performance following L-DOPA treatment, and sought to identify brain sites through which these receptors were acting. We first administered M1R-preferring antagonist trihexyphenidyl (0, 0.1, and 1.0 mg/kg, i.p.) or the M4R-preferring antagonist tropicamide (0, 10, and 30 mg/kg, i.p.) before L-DOPA, after which LID and motor performance were evaluated. Both compounds worsened and extended the time course of LID, while M1R blockade improved motor performance. We then evaluated the effects of tropicamide and trihexyphenidyl on dyskinesia induced by D1R agonist SKF81297 or D2R agonist quinpirole. Surprisingly, both M1R and M4R antagonists reduced D1R agonist-induced dyskinesia but not D2R agonist-induced dyskinesia, suggesting that mAChR blockade differentially affects MSN firing in the absence of postsynaptic DA. Finally, we evaluated effects of striatum- or PPN-targeted tropicamide microinfusion on LID and motor performance. Despite prior evidence, M4R blockade in either site alone did not affect the severity of LID via local striatal or PPN infusions. Taken together, these data suggest M4R as a promising therapeutic target for reducing LID using more selective compounds.

Blockade of muscarinic acetylcholine receptors facilitates motivated behaviour and rescues a model of antipsychotic-induced amotivation.

Disruptions to motivated behaviour are a highly prevalent and severe symptom in a number of neuropsychiatric and neurodegenerative disorders. Current treatment options for these disorders have little or no effect upon motivational impairments. We assessed the contribution of muscarinic acetylcholine receptors to motivated behaviour in mice, as a novel pharmacological target for motivational impairments. Touchscreen progressive ratio (PR) performance was facilitated by the nonselective muscarinic receptor antagonist scopolamine as well as the more subtype-selective antagonists biperiden (M1) and tropicamide (M4). However, scopolamine and tropicamide also produced increases in non-specific activity levels, whereas biperiden did not. A series of control tests suggests the effects of the mAChR antagonists were sensitive to changes in reward value and not driven by changes in satiety, motor fatigue, appetite or perseveration. Subsequently, a sub-effective dose of biperiden was able to facilitate the effects of amphetamine upon PR performance, suggesting an ability to enhance dopaminergic function. Both biperiden and scopolamine were also able to reverse a haloperidol-induced deficit in PR performance, however only biperiden was able to rescue the deficit in effort-related choice (ERC) performance. Taken together, these data suggest that the M1 mAChR may be a novel target for the pharmacological enhancement of effort exertion and consequent rescue of motivational impairments. Conversely, M4 receptors may inadvertently modulate effort exertion through regulation of general locomotor activity levels.

Of molecules, memories and migration: M1 acetylcholine receptors facilitate spatial memory formation and recall during migratory navigation.

Many animals use complex cognitive processes, including the formation and recall of memories, for successful navigation. However, the developmental and neurological processes underlying these cognitive aspects of navigation are poorly understood. To address the importance of the formation and recollection of memories during navigation, we pharmacologically manipulated turtles (Chrysemys picta) that navigate long distances using precise, complex paths learned during a juvenile critical period. We treated freely navigating turtles both within and outside of their critical learning period with a specific M1 acetylcholine receptor antagonist, a drug known to disrupt spatial cognition. Experienced adult turtles lost all navigational ability under the influence of the drug, while naive juveniles navigated successfully. We retested these same juveniles the following year (after they had passed their critical period). The juveniles that initially navigated successfully under the influence of the antagonist (but were unable to form spatial memories) were unable to do so subsequently. However, the control animals (who had the opportunity to form memories previously) exhibited typical navigational precision. These results suggest that the formation of spatial memories for navigation occur during a critical period, and successful navigation after the critical period is dependent upon the recall of such memories.

Muscarinic acetylcholine receptor M1 mediates prostate cancer cell migration and invasion through hedgehog signaling.

The autonomic nervous system contributes to prostate cancer proliferation and metastasis. However, the exact molecular mechanism remains unclear. In this study, muscarinic acetylcholine receptor M1 (CHRM1) expression was measured via immunohistochemical analysis in human prostate cancer tissue array slides. PC-3, LNCaP, and A549 cells were treated with pirenzepine or carbachol, and the cell migration and invasion abilities were evaluated. Western blotting and quantitative real-time PCR were performed to measure GLI family zinc finger 1 (GLI1), patched 1 (PTCH1), and sonic hedgehog (SHH) expression levels. High expression of CHRM1 was found in early-stage human prostate cancer tissues. In addition, the selective CHRM1 antagonist pirenzepine inhibited PC-3, LNCaP, and A549 cell migration and invasion, but the agonist carbachol promoted the migration and invasion of these three cell lines. Muscarinic signaling can be relayed by hedgehog signaling. These data show that CHRM1 is involved in the regulation of prostate cancer migration and invasion through the hedgehog signaling pathway.