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. Author manuscript; available in PMC: 2020 Aug 25.
Published in final edited form as: Behav Pharmacol. 2020 Jun;31(4):359–367. doi: 10.1097/FBP.0000000000000537

Comparison of the muscarinic antagonist effects of scopolamine and L-687,306

Gail Winger a, Emily M Jutkiewicz b, James H Woods a
PMCID: PMC7446609  NIHMSID: NIHMS1612821  PMID: 31922966

Abstract

This study aimed to use central and peripheral assays to compare the effects of the muscarinic antagonist scopolamine with those of a novel muscarinic antagonist, L-687,306 [(3R,4R)-3-(3-cyclopropyl-1,2,4,oxadiazol[5-yl]-1-azabicyclo[2.2.1]heptane. Groups of rats were trained to discriminate the stimulus effects of the muscarinic agonist, arecoline (1.0 mg/kg); concomitant measures of response rate were recorded. Separate groups were prepared with telemetery devices for recording bradycardia induced by arecoline (10 mg/kg). Methyl arecoline and arecoline were nearly equally potent in producing a brief but profound bradycardia, indicative of an equivalent effect in the heart. L-687,306 and scopolamine were both able to block this peripheral effect of arecoline. L-687,306 produced a surmountable antagonism of both the discriminative and rate-suppressing effects of arecoline. Scopolamine, however, was unable to antagonize the rate-reducing effects of arecoline in the discrimination assay. This limited the number of rats that could respond to the discriminative stimulus effects of arecoline, as well as the amount of arecoline stimulus effects they were able to report. The data suggest that L-687,306 may be a more generally effective muscarinic antagonist than scopolamine and support earlier reports that this antagonist has less direct effect on behavior.

Keywords: arecoline; discriminative stimulus; heart rate; L-687,306; methyl arecoline; scopolamine

Introduction

Although it might be assumed that drugs that have a common action of blocking central muscarinic receptors would produce common behavioral outcomes, this has not consistently been found to be the case. Witkin et al. (1987), for example, measuring rates of responding in rats performing a fixed ratio 10, food-reinforced task, found that some antimuscarinic compounds (benactyzine, aprophen, and adiphenine) increased rates of responding at low doses, whereas others (azaprophen, atropine, and pirenzepine) only decreased rates of responding. The rank-order potency of these drugs also differed in three in vitro assays, suggesting interactions with different receptor subtypes in the various assays. Grauer and Kapon (1996) evaluated relatively small doses of five muscarinic antagonists on a task that alternated between a visual discrimination task, thought to require primarily reference memory, and a spatial discrimination task, thought to involve working memory. Each of the antagonists (aprophen, scopolamine, atropine, benactyzine, and artane) produced impairment in the working memory task. Atropine, however, suppressed general behavior more than the other drugs, whereas aprophen had the least effects on general behavior and memory. Genovese et al. (1990) also noted that aprophen, although less potent than scopolamine and atropine, produced less behavioral suppression and was an effective antagonist of physostigmine across a wider dose range than the other two muscarinic antagonists.

Five distinct muscarinic receptors have been identified (Bonner et al., 1987; Caulfield, 1993), and differential binding to these receptors may contribute to the differences observed among the behavioral effects of antagonists. The strong structural homology among the receptors, however, has frustrated attempts to synthesize selective muscarinic agonists or antagonists. More research attention has been paid to muscarinic agonists than to antagonists, largely because of evidence that dementia may involve a cholinergic deficit (e.g. Bartus et al., 1982) that might be mitigated by administration of muscarinic agonists. Muscarinic antagonists, particularly scopolamine, are often given to produce those deficits in animal models of dementia, as well as in human volunteers (e.g. Drachman, 1977; Ebert and Kirch, 1988). L-687,306 was one in a series of compounds that was developed some time ago in an attempt to identify cholinergic agonist cognitive enhancers that had a reduced side-effects profile by virtue of having reduced efficacy and more selective subtype affinity (Freedman et al., 1992; Freedman et al., 1993). L-687,306 had no efficacy at M2 and M3 receptor subtypes and it produced no agonist action in autonomic measures (no salivation, hyperthermia, diarrhea, or heart rate changes) (Freedman et al., 1993) and no suppression of operant food-reinforced responding in rodents (Dawson and Iversen, 1993) at doses up to 30 mg/kg. L-687,306 had very limited efficacy at the M1 receptor subtype as measured in peripheral nervous system functional assays. Interestingly, it was able to reverse much of scopolamine’s disruptive effects in several memory tasks (Dawson and Iversen, 1993; Freedman et al., 1993). Although the authors did not suggest a mechanism of this scopolamine antagonist action, the tone of the article indicated that they would likely ascribe this antagonism to the limited efficacy of L-687,306 at M1 receptor subtypes. In our studies, we regarded and evaluated L-687,306 as a nonselective muscarinic antagonist; its efficacy at M1 receptors seems too slight to warrant anything but antagonism, even at this site.

In the work described here, the ability of scopolamine, L-687,306, and N-methylscopolamine (NMS) to antagonize the effects of the nonselective muscarinic agonist arecoline was evaluated using a centrally mediated assay (drug discrimination and concomitant suppression of ongoing responding) and a peripherally preferring assay (heart rate). Drug discrimination procedures have been used successfully to study agonist–antagonist interactions of drugs mediated through a number of receptors (e.g. Bertalmio and Woods, 1987; Jutkiewicz et al., 2011; Zanettini et al., 2014). Although these procedures are most revealing when at least some of the compounds evaluated have established interactions with a specific receptor or receptor subtype, they can be useful even in circumstances where receptor-selective effects are not simple to discern.

The effects of several doses of arecoline and methyl arecoline on heart rate of intact, freely moving rats were evaluated in the peripheral assay, and antagonism by the three nonselective antagonists was studied. The heart is thought to have primarily M2 receptors (Harvey, 2012), which mediate an M-2 agonist-induced decrease in rate. Because these measures were taken in freely moving, intact rats, other cholinergic influences on heart rate such as blood pressure could modify this response.

Methods

Cardiovascular assay

The cardiovascular procedures are described in detail in Jutkiewicz et al. (2013), and follow the description of assays by Delaunois et al. (2009).

Subjects

Male Sprague–Dawley rats, purchased from Harlan, Inc. (Indianapolis, Indiana) were housed in groups of three with food and water freely available. Housing and experimental rooms were maintained on a 12-h light/dark cycle with lights on at 7:00 a.m., and an average temperature of 21°C. Experiments described below occurred during the light cycle, between 10:00 a.m. and 3:00 p.m. Experimental protocols were approved by the University of Michigan University Committee on the Use and Care of Animals and the University of Texas Health San Antonio Institutional Animal Care and Use Committees, and conformed to the guidelines established by the NIH Guide for the Use of Laboratory Animals.

Surgical procedures

To measure changes in heart rate and mean arterial pressure (MAP), rats were implanted with telemetric transmitters (TA11PA-C40 or TL11M2-C50-PXT, Data Sciences International, Transoma Medical Inc., St. Paul, Minnesota, USA) under ketamine (90 mg/kg, i.p.) and xylazine (10 mg/kg, i.p.) anesthesia. Each transmitter was placed into a subcutaneous pocket on the side of the abdomen, and the catheter extending from the base of the transmitter was inserted 2–3 cm into the femoral artery and secured with a suture. Following surgery, rats were singly-housed and allowed to recover for at least 7 days before experimentation. All rats continued to have free access to food and water at all times.

Telemetry system

The system consisted of battery-operated subcutaneous transmitters, Physiotel receivers, the DSI Data Exchange Matrix, and the Dataquest A.R.T. system. These transmitted, collected and stored the digital data on blood pressure and heart rate to a computer (Data Sciences International, Transoma Medical Inc., St. Paul, Minnesota, USA). Blood pressure and heart rate data were compiled by the Dataquest A.R.T. Gold Analysis 3.01 software.

Experimental design

At the start of an experimental session, rats’ home cages were placed on top of the receivers, and data were collected for at least 1 h from undisturbed animals. Each rat was then given a saline injection to provide initial cardiovascular responses to the injection and handling procedures (saline habituation). Thirty minutes later a second injection was administered, typically either a second saline injection or the designated pretreatment drug (pretreatment). Fifteen minutes later, either saline or the test drug was given as the final injection (treatment). Data were collected for at least 2 h following the last injection. All injections were administered subcutaneously (s.c.) in a volume of 1 ml/kg.

Each rat was used to evaluate multiple experimental conditions; each rat received four to six different treatments, one per week with at least 6 days between drug exposures.

Data analysis

The analysis program calculated an average heart rate and MAP every 10 s. These 10-s epochs were subsequently averaged over 1 min per rat, and data from four to six rats (except as noted below) were averaged for each treatment group with SEM as a measure of variability.

The graph of each antagonist shows data for 20 min following the last (treatment) injection, which was of 10 mg/kg arecoline 15 min following administration of an antagonist. The control data with saline were taken from an historical average (N = 17) as were the control data with 10 mg/kg arecoline alone (N = 18).

Drug discrimination assay

The drug discrimination procedures are described in detail in Jutkiewicz et al. (2011).

Subjects

Male Sprague-Dawley rats were purchased from Harlan (Indianapolis, Indiana) and singly housed in polycarbonate cages with water continuously available. All rats weighed approximately 280–290 g at the start of the experiment. Housing and experimental rooms were maintained on a 12-h light/dark cycle with lights on at 7:00 a.m. and an average temperature of 21°C. The rats’ weights were allowed to increase gradually and were maintained at approximately at 350–375 gm with a food-restricted diet of Purina rodent food. A group of six rats was trained to discriminate 1 mg/kg arecoline and evaluated in the described experiments.

Apparatus

Drug discrimination procedures were performed in six standard operant conditioning chambers with an area of 30.5 × 24.1 × 21.0 cm and stainless steel grid floors (ENV-008; Med Associates, St. Albans, Vermont) contained within ventilated, sound-attenuating boxes. Each chamber was equipped with two nose-poke devices with apertures containing yellow LED lights on either side of a dipper capable of delivering 50 μl of fluid into a third opening (H14–05R; Coulbourn Instruments, Whitehall, Pennsylvania).

Discrimination training

All subjects were initially trained to respond on a fixed-ratio (FR) 10 schedule of reinforcement in either aperture. Completion of the schedule requirements resulted in 10-s access to 50 μl of vanilla-flavored Ensure (Abbott Laboratories, Abbott Park, Illinois). Side or aperture preferences were determined, and the saline- or arecoline-associated nose-poke apertures were assigned to each rat in the following format to minimize biases: (1) approximately 50% of rats had the drug-associated aperture assigned on the left and 50% of the rats had the drug-associated aperture assigned on the right; and (2) the drug-associated aperture was assigned to the nonpreferred side in 50% of subjects.

Arecoline (1.0 mg/kg) was administered to a rat just before it was placed in the chamber, and 5 min later, a house light and both apertures were illuminated. One of the apertures was designated ‘correct’ following drug administration, and responses in that aperture resulted in delivery of liquid reinforcement. When saline rather than arecoline was injected before a session, the other aperture was designated as ‘correct’; responses in the ‘correct’ aperture produced the reinforcer.

Rats were considered ready for testing when the following criteria were met on three consecutive days of training: (1) responding on first FR of the session was completed on the injection-appropriate aperture; and (2) >85% of total session responses were made on the injection-appropriate aperture.

Discrimination testing and maintenance

During test sessions, completed FRs in either nose-poke aperture were reinforced with 10-s access to Ensure. For evaluation of antagonist effects, a dose of the selected antagonist was administered 15 min before the start of the session, and animals were returned to their home cages. Arecoline was then given immediately before the rats were placed in the response chambers as indicated earlier. Rats received no more than two test sessions per week; at least once per month dose-effect curves with arecoline were determined to monitor for changes in drug sensitivity. Full or complete generalization to a discriminative cue was defined as >85% of responding on the drug-associated aperture and completing at least one FR.

Data analysis

The data for discrimination dose-effect curves are averaged over five to six rats. Discrimination data are expressed as a percentage of responses occurring on the arecoline-associated aperture out of the total number of responses on both the drug- and saline-associated apertures. Rates of responding were calculated by dividing the total number of responses by the total duration of SD presentations. Data were averaged across five to six rats in every dose condition unless otherwise described under the section ‘Results’. Response rates were included in group averages only if at least one FR was completed.

Drugs

Drugs (−) scopolamine hydrochloride and (−) scopolamine methyl bromide (NMS) were purchased from Sigma Aldrich, St. Louis, Missouri. Arecoline hydrobromide was purchased from Acros Organics. Methylarecoline was a gift from Dr. E.F. Domino. L-687,306 was generously donated by Merck Pharmaceuticals, Kenilworth, New Jersey. All drugs were dissolved in sterile saline before administration and were given as the salt form.

Results

Cardiovascular assay

Figure 1 (top) shows the effects of saline pretreatment followed by each of three doses of arecoline on heart rate. The heart rate increase that is an artifact of the injection procedure is shown at the earliest time points. Recall that the saline administrations shown here are the second injections for each rat; saline habituation injections were given 30 min before the saline pretreatment injections shown in the figure. Arecoline produced dose-related decreases in heart rate; these decreases were short-lived, with rates returning to near base-line levels at about 10 min following the injection.

Fig. 1.

Fig. 1

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Effects of i.p. administration of saline (open symbols, min 0–15) and arecoline (top) or methyarecoline (bottom) (closed symbols, min 15–40) on heart rate in beats per minute. Symbols (circle, square, triangle) indicate test conditions consisting of a saline injection at time 0 (open symbols) followed 15 min later by one of the three doses of arecoline (top graph; filled symbols), or one of the two doses of methylarecoline (bottom graph; filled symbols). All data points are an average of eight rats with variability indicated as SEM.

The data in Fig. 1 (bottom) represent heart rate measures after saline pretreatment and following administration of 10 or 18 mg/kg methylarecoline. Methylarecoline, like arecoline, produced a dose-related decrease in heart rate following administration of these doses; this quaternary form of arecoline was equally effective and slightly less potent than arecoline itself. A dose of 32 mg/kg of methylarecoline was lethal in the rats.

Figure 2 shows the ability of several doses of scopolamine (a), NMS (b), and L-687,306 (c) to prevent the bradycardia produced by 10 mg/kg arecoline. There was a dose-related ability of scopolamine (Fig. 2a) to prevent arecoline’s effects on heart rate, with complete antagonism observed following a dose of 0.1 mg/kg. NMS also reversed arecoline’s bradycardiac effects and was 10 times more potent than scopolamine in this regard; a dose of 0.01 mg/kg of NMS completely antagonized the effects of arecoline on heart rate and appeared to produce some ‘overshoot’ tachycardia (Fig. 2b). L-687,306 was 10 times less potent than scopolamine in reversing arecoline’s effects on heart rate, as shown in Fig. 2c. A dose of 1.0 mg/kg L-687,306 completely reversed the effects of 10 mg/kg arecoline on heart rate.

Fig. 2.

Fig. 2

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Effects of saline or three doses of scopolamine (a), N-methylscopolamine (NMS) (b), and L-687,306 (c) on arecoline-induced decreases in heart rate. Arecoline (black circles) was given at time 0, which was 30 min after a saline habituation injection (not shown), and 15 min after injection of saline or an antagonist at the indicated dose. Open circles show the effects of saline, and decreasingly dark circles show the effects of increasing doses of each of the three antagonists. All data points are an average of eight rats with variability indicated as SEM.

Drug discrimination

A dose-effect curve for arecoline in the arecoline discrimination assay is shown on the top part of Fig. 3. Figure 3 also demonstrates that arecoline methyl iodide was unable to mimic the discriminative stimulus effects of arecoline (top), and was nearly 10 times less potent suppressing rates of responding (bottom). The effects of NMS on arecoline’s discriminative stimulus and rate-suppressing effects were evaluated as well; NMS had no effect on either of arecoline’s behavioral effects (data not shown).

Fig. 3.

Fig. 3

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Antagonism by three doses of L-687,306 of the discriminative stimulus and rate-suppressing effects of arecoline. Abscissae: dose of arecoline in mg/kg. Ordinate (top) percent responding on the arecoline-appropriate lever. Ordinate (bottom) rate of responding in responses/s. Data points are the means of five (+0.03 mg/kg L-687,306), six (arecoline alone, + 0.01 L-687,306; 1.0 L-687,306), or seven (+ 0.1 mg/kg L-687,306) rats. Vertical lines through data points represent SEM and are not shown when this measure of variability is contained within the data point.

Figure 4 shows the effects of 0.01, 0.1 and 1.0 mg/kg L-687,306 on the arecoline discrimination. The bottom of Fig. 4 shows the interaction of these doses of L-687,306 and arecoline on rates of responding in the discrimination assay. This muscarinic antagonist produced a dose-related rightward shift in both the stimulus and rate reducing effects of the muscarinic agonist. A dose of 0.01 mg/kg L-687,306 shifted both arecoline dose-response curves approximately ¼ log unit to the right; a dose of 0.1 mg/kg L-687,306 shifted arecoline a further ¼ log unit to the right; a dose of 1.0 mg/kg L-687,306 completely abolished both the discriminative stimulus and rate-suppressing effects of arecoline up to 5.6 mg/kg.

Fig. 4.

Fig. 4

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Effects of increasing doses of arecoline on arecoline-appropriate responding (top) and rates of responding (bottom) (solid circles), and the ability of three doses of scopolamine to modify these behaviors. Data points are the means of five (+ 0.01 scopolamine) or six (arecoline alone) rats, except where noted in the figure. N = 3 and N = 2 in the figure indicate that behavior was sufficiently suppressed in the remaining three or four rats to preclude including them in the figure. Vertical lines through data points represent SEM and are not shown when this measure of variability is contained within the data point.

Figure 5 presents the interaction of scopolamine and arecoline in the arecoline-discrimination assay. Although the largest dose of scopolamine (0.1 mg/kg) tended to produce a rightward shift of 3/4 log unit in arecoline’s discriminative stimulus effects, the maximum percent arecoline-appropriate responding following this dose was no greater than 50% on average, and data from only three rats could be evaluated because their rates of responding were too low at this or smaller doses of scopolamine + arecoline.

Fig. 5.

Fig. 5

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Inability of arecoline methyl iodide to produce arecoline-like stimulus effects at doses that suppressed rates of responding. Data points are the mean of six rats for arecoline and the mean of four rats for arecoline methyl iodide. Vertical lines through data points represent SEM and are not shown when this measure of variability is contained within the data point.

Larger doses of arecoline could not be tested because this combination of scopolamine and arecoline dramatically reduced rates of responding. These data indicate that arecoline’s discriminative stimulus effects may be at least somewhat reversed by scopolamine, but the rate-decreasing effects of arecoline are insensitive to scopolamine’s antagonist effects. Very similar interactions were shown in attempts to antagonize the discriminative stimulus effects of arecoline with atropine (data not shown).

Discussion

Arecoline demonstrated both central and peripheral effects in these assays. Peripheral effects were demonstrated by bradycardia that was produced by both arecoline and N-methyl arecoline, largely peripherally restricted analog, and was blocked by the largely peripherally restricted antagonist NMS. Central effects of arecoline were identified in the discriminative stimulus effects of the drug. The central action was indicated by the finding that N-methyl arecoline did not have discriminative stimulus effects in common with arecoline. N-methyl arecoline did suppress rates of responding at doses that were close to those that produced lethality. Arecoline-induced suppression of response rates could therefore include a peripheral mechanism as has been suggested by Witkin (1989), who found that centrally and peripherally acting muscarinic agonists were both able to suppress avoidance responding in squirrel monkeys in an apparently redundant manner.

Scopolamine and L-687,306 are muscarinic antagonists with a similar profile of receptor binding interaction insofar as they have been studied (Freedman et al., 1993). The data presented here confirm their similarity in measures of peripheral antimuscarinic effects. Both scopolamine and L-687,306, as well as N-methyl scopolamine, produced a dose-related antagonism of the profound bradycardia that followed administration of 10 mg/kg arecoline. However, the two antagonists were not similar with respect to their ability to antagonize the largely central effects of arecoline. Although L-687,306 produced an initially surmountable, and eventually an apparently unsurmountable blockade of both the discriminative stimulus effects and the rate suppressing effects of arecoline, scopolamine’s ability to modify the central effects of arecoline was more limited. As has been found by others (Jung et al., 1987), scopolamine was unable to attenuate the rate-suppressing effects of arecoline, whether these were centrally or peripherally mediated or both. This limited the sample size for evaluation of antagonism of the discriminative stimulus effects of arecoline. The few animals that did respond, albeit at low rates following scopolamine administration, showed a rightward but incomplete shift in the arecoline dose-response curve. It is possible that scopolamine has rate-suppressing effects of its own, which L-687,306 lacks, which prevented the older compound from antagonizing the rate suppressing effects of arecoline. Interestingly, however, a dose of 0.1 mg/kg scopolamine, which suppressed rates of responding dramatically when combined with arecoline (Fig. 5), produced only modest decreases in rates of responding when given alone (unpublished observations). In contrast, a dose of 3 mg/kg L-687,306, three times larger than the dose that blocked the discriminative and rate-decreasing effects of arecoline, had no effect on response rates when given alone (Woods, unpublished observations), and Dawson and Iversen (1993) reported that L-687,306 did not impair chain-pulling behavior in rats at doses up to 30 mg/kg.

As mentioned in the introduction, aprophen is a muscarinic antagonist that appears to produce less direct behavioral suppression than other muscarinic antagonists (Witkin et al., 1987; Genovese et al., 1990; Grauer and Kapon, 1996). As noted by Witkin et al. (1987), some drugs ‘may be effective antimuscarinic agents in vivo at doses that do not produce the spectrum of undesirable effects (e.g. behavioral) found with existing centrally active antimuscarinic compounds’ (page 802). L-687,306 appears to be one such central muscarinic antagonist. It will be interesting to determine whether the behavioral suppressant effects of classic muscarinic antagonists such as scopolamine and atropine are mediated through activity at different sets of muscarinic receptors than those that mediate the effects of L-687,306 and aprophen, or through activity at nonmuscarinic receptors. Nevertheless, taken together, these data suggest that it may be possible to establish a pharmacological block of the muscarinic nervous system without dramatically compromising ongoing behavior. This has important applications for eventual clinical and experimental application of antimuscarinic drugs and general understanding of muscarinic activity.

The cholinergic nervous system, particularly the muscarinic aspect of this system, has important implications in at least three human afflictions. (1) It has long been thought to have a likely role in dementia (e.g. Ferreira-Vieira et al., 2016; Kamkwalala and Newhouse, 2017). Scopolamine is the prototypical antimuscarinic compound used to produce cognitive deficits that serve as a model for human dementia (see review, Blokland et al., 2016). (2) It has recently been suggested as having involvement in depression (e.g. Dulawa and Janowsky, 2018), with the report that scopolamine may have antidepressant effects (Furey and Zarate, 2013; Park et al., 2019). (3) Blockade of the muscarinic cholinergic nervous system with the muscarinic antagonist atropine is the long-accepted, but less than ideal, treatment for organophosphate poisoning. It is important to evaluate novel antimuscarinic drugs in each of these conditions. If compounds such as L-687,306 and aprophen do not produce cognitive deficits of the type produced by scopolamine, we may gain greater understanding of the role of cholinergic circuitry and receptor mediation in cognitive decline. If they are able to relieve depression in some individuals, it may be possible to add antimuscarinics to the armamentarium against depression with less concern about increasing the risk of later dementia (Coupland et al., 2019). And they or chemically similar drugs may have advantages over atropine in treating organophosphate poisoning in that they can be given in larger doses with fewer side effects. In support of these possibilities, our preliminary data suggest that L-687,306, like scopolamine, produces positive effects in animal models of depression (Jutkiewicz, unpublished observations). In addition, it does not affect learning, attention, and memorial processes in nonclinical models in which scopolamine is disruptive (Kangas, unpublished observations). The development and study of novel cholinergic compounds is likely to have significant clinical usefulness.

Acknowledgements

The technical support of Jessica Priebe and Yong Gong Shi is greatly appreciated. We thank Dr. E.F. Domino for providing us a sample of quaternary arecoline. We acknowledge with sincere appreciation the efforts of Drs. Lisa Gold and Geoffrey Varty of Merck Pharmaceuticals to identify a source of and purity of L-687,306.

This study was supported by the Julian Villarreal Fund for Drug Abuse Research and RO1 MH107499 to J.H.W.

Footnotes

Conflicts of interest

There are no conflicts of interest.

References

  1. Bartus RT, Dean RL 3rd, Beer B, Lippa AS (1982). The cholinergic hypothesis of geriatric memory dysfunction. Science 217:408–414. [DOI] [PubMed] [Google Scholar]
  2. Bertalmio AJ, Woods JH (1987). Differentiation between mu and kappa receptor-mediated effects in opioid drug discrimination: apparent pa2 analysis. J Pharmacol Exp Ther 243:591–597. [PubMed] [Google Scholar]
  3. Blokland A, Sambeth A, Prickaerts J, Riedel WJ (2016). Why an M1 antagonist could be a more selective model for memory impairment than scopolamine. Front Neurol 7:167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bonner TI, Buckley NJ, Young AC, Brann MR (1987). Identification of a family of muscarinic acetylcholine receptor genes. Science 237:527–532. [DOI] [PubMed] [Google Scholar]
  5. Coupland CAC, Hill T, Dening T, Morriss R, Moore M, Hippisley-Cox J (2019). Anticholinergic drug exposure and the risk of dementia: a nested casecontrol study. JAMA Internal Med 179:1084–1093. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Caulfield MP (1993). Muscarinic receptors–characterization, coupling and function. Pharmacol Ther 58:319–379. [DOI] [PubMed] [Google Scholar]
  7. Dawson GR, Iversen SD (1993). The effects of novel cholinesterase inhibitors and selective muscarinic receptor agonists in tests of reference and working memory. Behav Brain Res 57:143–153. [DOI] [PubMed] [Google Scholar]
  8. Delaunois A, Dedoncker P, Hanon E, Guyaux M (2009). Repeated assessment of cardiovascular and respiratory functions using combined telemetry and whole-body plethysmography in the rat. J Pharmacol Toxicol Methods 60:117–129. [DOI] [PubMed] [Google Scholar]
  9. Drachman DA (1977). Memory and cognitive function in man: does the cholinergic system have a specific role? Neurology 27:783–790. [DOI] [PubMed] [Google Scholar]
  10. Dulawa SC, Janowsky DS (2019). Cholinergic regulation of mood: from basic and clinical studies to emerging therapeutics. Mol Psychiatry 24: 694–709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ebert U, Kirch W (1998). Scopolamine model of dementia: electroencephalogram findings and cognitive performance. Eur J Clin Invest 28:944–949. [DOI] [PubMed] [Google Scholar]
  12. Ferreira-Vieira TH, Guimaraes IM, Silva FR, Ribeiro FM (2016). Alzheimer’s disease: targeting the cholinergic system. Curr Neuropharmacol 14:101–115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Freedman SB, Patel S, Harley EA, Iversen LL, Baker R, Showell GA, et al. (1992). L-687,306: a functionally selective and potent muscarinic M1 receptor agonist. Eur J Pharmacol 215:135–136. [DOI] [PubMed] [Google Scholar]
  14. Freedman SB, Dawson GR, Iversen LL, Baker R, Hargreaves RJ (1993). The design of novel muscarinic partial agonists that have functional selectivity in pharmacological preparations in vitro and reduced side-effect profile in vivo. Life Sci 52:489–495. [DOI] [PubMed] [Google Scholar]
  15. Furey ML, Zarate CA Jr (2013). Pulsed intravenous administration of scopolamine produces rapid antidepressant effects and modest side effects. J Clin Psychiatry 74:850–851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Genovese RF, Elsmore TF, Witkin JM (1990). Relationship of the behavioral effects of aprophen, atropine and scopolamine to antagonism of the behavioral effects of physostigmine. Pharmacol Biochem Behav 37:117–122. [DOI] [PubMed] [Google Scholar]
  17. Grauer E, Kapon J (1996). Differential effects of anticholinergic drugs on paired discrimination performance. Pharmacol Biochem Behav 53:463–467. [DOI] [PubMed] [Google Scholar]
  18. Harvey RD (2012). Muscarinic receptor agonists and antagonists: Effects on cardiovascular function In: Fryer AD, Christopoulos A, Mathanson NM, editors. Muscarinic Receptors. Berlin: Springer-Verlag; pp. 299–316. [DOI] [PubMed] [Google Scholar]
  19. Jung M, Costa L, Shearman GT, Kelly PH (1987). Discriminative stimulus properties of muscarinic agonists. Psychopharmacology (Berl) 93:139–145. [DOI] [PubMed] [Google Scholar]
  20. Jutkiewicz EM, Rice KC, Carroll FI, Woods JH (2013). Patterns of nicotinic receptor antagonism II: cardiovascular effects in rats. Drug Alcohol Depend 131:284–297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jutkiewicz EM, Brooks EA, Kynaston AD, Rice KC, Woods JH (2011). Patterns of nicotinic receptor antagonism: nicotine discrimination studies. J Pharmacol Exp Ther 339:194–202. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kamkwalala AR, Newhouse PA (2017). Beyond acetylcholinesterase inhibitors: novel cholinergic treatments for Alzheimer’s disease. Curr Alzheimer Res 14:377–392. [DOI] [PubMed] [Google Scholar]
  23. Klinkenberg I, Blokland A (2010). The validity of scopolamine as a pharmacological model for cognitive impairment: a review of animal behavioral studies. Neurosci Biobehav Rev 34:1307–1350. [DOI] [PubMed] [Google Scholar]
  24. Moore H, Dudchenko P, Comer KS, Bruno JP, Sarter M (1992). Central versus peripheral effects of muscarinic antagonists: the limitations of quaternary ammonium derivatives. Psychopharmacology (Berl) 108:241–243. [DOI] [PubMed] [Google Scholar]
  25. Park L, Furey M, Nugent AC, Farmer C, Ellis J, Szczepanik J, et al. (2019). Neurophysiological changes associated with antidepressant response to ketamine not observed in a negative trial of scopolamine in major depressive disorder. Int J Neuropsychopharmacol 22:10–18. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Witkin JM, Gordon RK, Chiang PK (1987). Comparison of in vitro actions with behavioral effects of antimuscarinic agents. J Pharmacol Exp Therap 242:796–803. [PubMed] [Google Scholar]
  27. Witkin JM (1989). Central and peripheral muscarinic actions of physostigmine and oxotremorine on avoidance responding of squirrel monkeys. Psychopharmacology (Berlin) 97:376–382. [DOI] [PubMed] [Google Scholar]
  28. Zanettini C, France CP, Gerak LR (2014). Quantitative pharmacological analyses of the interaction between flumazenil and midazolam in monkeys discriminating midazolam: determination of the functional half life of flumazenil. Eur J Pharmacol 723:405–409. [DOI] [PMC free article] [PubMed] [Google Scholar]

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