Brit. J. Phannacol. (1954), 9, 218.

A COMPARISON OF THE PERIPHERAL PARASYMPATHOLYTICAND AUTONOMIC GANGLION BLOCKING ACTIVITIES

OF METHANTHELINE (" BANTHINE ") ANDPROPANTHELINE (" PRO-BANTHINE ") WITH

ATROPINE AND HEXAMETHONIUMBY

E. A. JOHNSON AND D. R. WOODFrom the Department of Pharmacology and Therapeutics, University of Sheffield

(RECEIVED MARCH 6, 1954)

Originally prepared by Burtner and Cusic (1943),methantheline (,8-diethylaminoethyl xanthene-9-carboxylate methobromide) has been studiedpharmacologically by several workers, notablyHambourger, Cook, Winbury, and Freese (1950),Lehman and Knoeffel (1944), Chittumn, Longino,Metcalf, and Grimson (1949), Longino, Grimson,Chittum, and Metcalf (1950), and Lyons, Reeves,and Grimson (1950). Their work shows that withprogressively increasing doses and concentrationsof methantheline there appears first an atropine-like action, followed, at a higher dosage range, byautonomic ganglion blockade and finally, in rela-tively high doses, by a curare-like action.

Propantheline (2'-diisopropylaminoethyl xan-thene-9-carboxylate methobromide) is the iso-propyl analogue of methantheline and differsfrom it only in degree of activity (Hambourger,1952), its actions being qualitatively similar.Although methantheline and propantheline seem

to have considerable atropine-like activity, therehave been few comparative studies with atropinein animals. It is the purpose of the present workto provide such a study. At the same time a com-parison of the autonomic ganglion blocking activi-ties, and of the curare-like activities, of the twodrugs has been made. The local anaesthetic activi-ties have also been investigated.

METHODSThe substances used were methantheline bromide,

propantheline bromide, hexamethonium bromide,atropine sulphate and acetylcholine chloride. Dosesand concentrations, and comparisons of activity, aregiven throughout this paper in terms of these salts.

Comparisons of Peripheral ParasympatholyticActivity

Isolated Guinea-pig Ileum.-Steady sub-maximalcontractions to ACh were obtained at two-minuteintervals. The dose of antagonist was given oneminute before the next dose of ACh. The antag-onistic effects of three separate doses of atropine,followed by three of methantheline, were measured bythe percentage reduction in height of the subsequentcontraction due to ACh.

Five such comparative assays of methantheline andatropine and five of propantheline and atropine wereperformed on pieces of gut from 10 guinea-pigs.The percentage reduction in height of contraction

was plotted against the logarithm of the dose of antag-onist; the potency of methantheline or propantheline,compared with atropine, was obtained as the mean ofthe ratios of doses required to produce 50% inhi-bition of the height of contraction.Blood Pressure in the Cat.-In each experiment a

suitable dose of ACh was discovered (0.1 to 1.0 pg.)which would give an adequate fall in blood pressure.Comparisons of activity were made by relating thedose of each antagonist needed to inhibit to an equaldegree the response to ACh.

Mydriatic Activity in the Mouse.-The method ofIng, Dawes and Wajda (1945) was used.

Intraperitoneal injections of atropine (2, 4, and 8ug. in 0.2 ml. 0.9% NaCI) were made into three groupsof five mice and the pupil diameter measured 15min. later by viewing the eye under a strong lightwith the aid of a binocular microscope provided witha scale in the eyepiece. This procedure was repeatedtwice on the same day in six further groups of fivemice, so that 45 animals in all were used. Three dosesof methantheline (10, 20, and 40 Mg., or 5, 10, and20 pg.) were investigated in another 45 mice on thesame day; four such complete experiments were per-formed. Similarly three complete experiments with

METHANTHELINE AND PROPANTHELINE

4, 8, and 16 pg. doses of propantheline, and 2, 4, and8 pg. doses of atropine, were performed.The results from the four experiments on methan-

theline and the three on propantheline were pooledseparately and subjected to a suitable form ofstatistical analysis for calculating the regression lineof the dose of mydriatic on mean pupil diameter(Emmens, 1948). From the two regression lines formethantheline and atropine, and the two for pro-pantheline and atropine, the comparisons of potencywere made.

Information about the duration of mydriatic acti-vity was obtained by plotting curves relating meanpupil size to the time after intraperitoneal injection ofthe equiactive doses of 40 pg., 16 pg., and 8 pg. ofmethantheline, propantheline, and atropine respec-tively. Groups of 15 mice received atropine andmethantheline in one experiment, and atropine andpropantheline in another.

Salivary Secretion in the Cat.-The method usedwas that of Btilbring and Dawes (1945). A steadyflow of saliva was produced by an intravenousinfusion at a uniform rate (0.5 ml./min.) of 0.004%(w/v) of carbachol in Ringer-Locke solution. Adren-aline hydrochloride (0.002%) was included to offset tosome extent the fall in blood pressure produced bythe carbachol. The rate of secretion was recorded bya drop timer.The relative activity of methantheline or propan-

theline and atropine was found by recording twoeffects of a dose of one substance between two effectsof a dose of the other. This procedure was repeateduntil an exact match was obtained, or until two dosesof one drug were found more and less effective respec-tively than a fixed dose of the other. The activity wasjudged by the maximum effect of each injection. Fivecats were used for each assay. In each of the tencats a comparison was made at several dose levels.In eight of the cats the activities of methantheline andatropine or propantheline and atropine in inhibitingthe carbachol-induced fall in blood pressure wererecorded.

Comparisons of Ganglion Blocking ActivityNictitating Membrane of the Cat.-The autonomic

ganglion blocking property of methantheline bromidewas compared with that of hexamethonium in fourcats anaesthetized with pentobarbitone sodium, andthat of propantheline with hexamethonium in afurther four cats, by relating the ability of the drugsto reduce the recorded height of contraction of thenictitating membrane in response to pre-ganglionicnerve stimulation. Stimulation was by square waveshocks, of 800 psec. duration and 45 c/sec., automatic-ally applied, every 2 min. for 3 to 10 sec., to thecentral end of the cut vago-sympathetic trunk.Steady contractions were obtained and the drug wasgiven by vein at a fixed time before the subsequentcontraction. The reduction in height of the con-traction following the drug was expressed as a per-centage of the previous contraction height and was

plotted against the logarithm of the dose/kg. for eachdrug. Because the lines were not parallel, the relativepotency was obtained by comparing the two log. dose-response lines for each pair of drugs at variousresponse levels.Blood Pressure in the Cat.-The relative ability of

methantheline and hexamethonium to lower theblood pressure was observed in four cats, and that ofpropantheline and hexamethonium in three. All thecats were anaesthetized with pentobarbitone sodium.

Acetylcholine Hypertension in the AtropinizedCat.-In four cats anaesthetized with pentobarbitonesodium a comparison was made of the relative powersof hexamethonium, methantheline, and propanthelineto inhibit the rise in blood pressure caused by theganglion stimulation produced by large doses of ACh(0.1 to 0.2 mg./kg.) after large doses of atropine.Atropine (1 mg. /kg.) was given initially by vein.Several doses of methantheline or propantheline andof hexamethonium were tried in order to find one ofeach which would almost prevent the rise of bloodpressure following ACh.Comparisons of Curare-like ActivityCat Sciatic Nerve-Gastrocnemius Preparation.-

The cats were anaesthetized with pentobarbitonesodium. Drug injections were given into theproximal side of the ligatured ipsilateral external iliacartery. The sciatic nerve was stimulated by singlesupra-maximal thyratron 'shocks at 10 sec. intervals.In two cats the relative potency of propantheline andmethantheline was compared in producing inhibitionof the contraction of the gastrocnemius muscle.Comparisons of Local Anaesthetic Activity*An attempt was made to compare any local anaes-

thetic activity of methantheline with that of cocainehydrochloride by the guinea-pig method of Chanceand Lobstein (1944). Thelocalanaestheticwasappliedto both eyes and the corneal reflex tested 45 sec. laterand at every minute for 5 min. The number of timesthe reflex was present out of a possible maximum of10 for both eyes was recorded for several concen-trations of methantheline (5, 2.5 and 1%) and cocainehydrochloride (0.2%). The substances were dissolvedin 0.9% NaCl.Information regarding the duration, intensity, and

any evidence of irritant properties of methanthelineand propantheline as local anaesthetics was obtainedby limited experimentation in two human subjects.Injections of 0.2 ml. of solutions of methantheline,propantheline or procaine hydrochloride were madeinto the skin of the flexor surface of the forearm.The solutions were made up in 0.9% NaCl; controlinjections were of saline alone.

RESULTSPeripheral Parasympatholytic Activity

Guinea-pig Ileum.-The results of assays onguinea-pig ileum show propantheline to have amean potency (relative to atropine=l) of 1.95+

219

E. A. JOHNSON and D. R. WOOD

24 (35) (25)

18 (45)o _ (45

12(35

0(45) 0-O Atropine6 * Methantheline

I I I I I I I0 . 42 4 5 8 10Dose in pg./mouse

20 40

FIG. 1.-The log dose-response lines for methantheline bromide andatropine sulphate in producing mydriasis in the mouse. Ordinate:mean pupil diameter in arbitrary units of the microscope eye-piece scale. Abscissa: big. per mouse intraperitoneally. Thenumber of animals used for each point is indicated on the graph.The points shown on the graph are the observed mean values.The dose-response lines are calculated. The horizontal inter-rupted lines indicate the response levels at which the potencycomparisons were made.

0.08 S.E. (5 expts.) compared with 1.73±0.19 S.E.(5 expts.) for methantheline.

Intravenous Acetylcholine in the Cat.-In fourcats the ratio of activity of methantheline to atro-pine lay between 0.7 and 1.0; in a further fourcats the drugs were about equiactive.

Mydriasis in the Mouse.-The two log dose-response lines for methantheline and atropine areshown in Fig. 1 and those of propantheline andatropine in Fig. 2. The points shown on the graphare the observed mean values, whereas the dose-response lines are calculated.As can be seen, the log dose-response lines for

methantheline and atropine are far from parallel;it is therefore impossible to make an accurateestimate of the relative potency of the two drugs.An average has been taken of the relative poten-

24

E

0-12~~~

C O-O Atropine

6

OI I .I2 4 8 16Dose in pg./mouse

FIG. 2.-The log dose-response lines for propantheline bromide andatropine sulphate in producing mydriasis in the mouse (45 micefor each dose). Ordinate: mean pupil diameter in arbitraryunits of the microscope eyepiece scale. Abscissa: pg. permouse intraperitoneally. Points on graph are observed meanvalues. Dose-response lines are calculated.

cies calculated at four response levels, and theselevels are represented by the horizontal dottedlines between the regression lines. The regressionlines for propantheline and atropine, however, areparallel. The mean relative potency of methan-theline and atropine (=1) is 0.28 and the relativepotency of propantheline and atropine (= 1) is0.47.The durations of action of methantheline and

atropine are compared in Fig. 3 and of methan-theline and propantheline in Fig. 4. Doses ofdrugs used were those known to be equiactive at15 min. The time for half recovery to the pre-injection diameter for methantheline is much lessthan for atropine (45 min. and 120 min. for 40 jig.and 8 jug. respectively of methantheline and atro-pine). Propantheline, however, approached atro-pine in time for half recovery (67.5 min. and 77.5min. for 16 jug. and 8 jtg. respectively of prop-antheline and atropine).

O-o Atropine 8 pg./mouse0-0 Methantheline 40 pg./mouse

FIG. 3.-Duration of mydriasis in the mouse pro-duced by 40 pg. of methantheline bromide ismuch less than that following 8 pg. ofatropine sulphate intraperitoneally. Thesedoses were equiactive at 15 min. Ordinate:mean pupil diameter from 15 mice inarbitrary units of the microscope eyepiecescale. Abscissa: time after injection in min.

o

E£5-a0.

c0

220

I

METHANTHELINE AND PROPANTHELINE

FIG. 4.-Duration of mydriasis in the mouseproduced by 16 pg. of propantheline bromideis similar to that following 8 pg. of atropinesulphate intraperitoneally. These doses wereequiactive at 15 min. Ordinate: mean pupildiameter from 15 mice in arbitrary units ofthe microscope eyepiece scale. Abscissa: timeafter injection in min.

4-

ELO

-a0-

C

4c:

r-eo Atropine 8 pg./mouse-o Propantheline 16 pg.mouse

Minutes after Injection

Salivary Secretion.-The results of several com-parative assays of methantheline and atropine fromeach of five cats and those of several assays ofpropantheline and atropine in each of a furtherfive cats were averaged. The mean relative potencyof methantheline (relative to atropine= 1) calcu-lated from all the results is 2.23 ± 0.37 S.E. (5 expts.)and that for propantheline 2.95 ±0.41 S.E. (5expts.).

In inhibiting the hypotension produced by theinfusion of carbachol, methantheline and atropinewere approximately equipotent, whereas propan-theline had a mean potency of 2.7 (relative toatropine= 1).

Ganglion Blocking ActivityNictitating Membrane of the Cat.-The extent

of the fall in blood pressure produced by hexa-methonium was such that a dose which wouldcause over 50% inhibition of contraction couldnot be given in some cases without fear of killingthe animal. The log dose/kg.-response lines forthe action of methantheline and propantheline andhexamethonium on the nictitating membrane arenot parallel; estimates of relative potency were

2.5 mq aFIG. 5.-A typical record showing the effect of methantheline bromide

(M) in inhibiting the contraction of the nictitating membrane ofthe cat in response to pre-ganglionic sympathetic stimulationCat anaesthetized with pentobarbitone sodium. Time intervals,12 sec.

therefore made at several response levels and themean calculated for each experiment. Methan-theline has a mean potency (relative to hexa-methonium=l) in these experiments of 0.40±0.06S.E. (4 expts.) and propantheline (relative to hexa-methonium= 1) of 0.63 ± 0.08 S.E. (4 expts.) (Fig.5).

In two cats it was found that 5 mg. /kg. ofmethantheline or of propantheline had no effecton the contraction of the nictitating membraneproduced by intravenous adrenaline.

Reduction in Blood Pressure.-In three cats,0.5 mg. hexamethonium produced a fall in bloodpressure equal to that due to 5 mg. of methan-theline and in three other cats 5 mg. of propan-theline produced a fall in blood pressure equivalentto from 2.5 to 4 mg. of hexamethonium.

ACh Hypertension in the Atropinized Cat.Hexamethonium and methantheline were equi-active: propantheline was twice as active as hexa-methonium.

IM P

2 mg. 2 mg.

FIG. 6.-Record of the contractions of gastrocnemius muscle of thecat in response to stimulation of the sciatic nerve by singlesupramaximal shocks from a thyratron stimulator at a rate of5/min. The inhibitory effects of 2 mg. of propantheline bromide(P) and 2 mg. of methantheline bromide (M) given intra-arteriallyare seen to be equal. Cat anaesthetized with pentobarbitonesodium.

221

E. A. JOHNSON and D. R. WOOD

Curare-like ActivityPropantheline and methantheline showed curare-

like activity only in very large doses. They wereapproximately equiactive. The effects of 2 mg.of propantheline and of methantheline are shownin Fig. 6.

Local AnaesthesiaUsing three concentrations of methantheline

(5%, 2.5%, 1% in saline) instilled into the guinea-pig eye, no local anaesthetic activity could bedemonstrated in any of nine guinea-pigs. Com-pared with 0.2% cocaine hydrochloride, it pro-duced marked signs of irritation, especially withthe 5% and 2.5% concentrations. Similar resultswere obtained with propantheline in six guinea-pigs, except that there was some evidence of anaes-thesia with a 5 % concentration. As with methan-theline, marked signs of irritation were producedby the 2.5% and 5% solutions.

Experiments in man confirmed the irritant actionshown in the guinea-pigs. Intracutaneous injec-tions were followed by the development of aspreading weal and surrounding flare of vaso-

dilatation. Methantheline 1% in two subjectsproduced anaesthesia of slow onset but prolongedduration in the skin overlying the weal producedby the injection. Several minutes were neededfor the full development of anaesthesia, but itlasted for 6 to 7 hr. This is in contrast tothe immediate anaesthesia produced by 0.25%procaine, and which continues for only 25 to 30min. A higher concentration of methantheline,2%, produced anaesthesia of slightly more rapidonset and of much longer duration. There was

still some degree of anaesthesia 24 hr. later. Thelow concentration of 0.5% methantheline had a

similar effect to the 1% concentration. After 1and 2% methantheline, scarring and hypersensi-tivity of the skin were present for at least twoweeks. Exactly similar results were obtained withpropantheline, using 5%, 2.5%, and 1 % solu-tions. It is interesting to note that 0.1 c.c. ofeach of these solutions intradermally (equal toapproximately 8 mg.) in each subject producedcomplete arrest of salivary secretion and an in-creased pulse rate. The effect on salivary secretionlasted for approximately five hours. Inhibition ofgastro-intestinal activity was suggested by a feelingof fullness in the stomach, and loss of appetite.

DISCUSSIONThe present results with methantheline differ

considerably from those of other workers-notably Lehman and Knoeffel (1944). As far as

can be ascertained no systematic pharmacologicalexamination of the properties of propantheline hasyet been reported, apart from that of Hambourger(1952).Lehman and Knoeffel (1944) found methan-

theline to be 100 times weaker than atropine ininhibiting salivary secretion. Although their assaywas done on cats, their method was totally differentfrom that used here. They used pilocarpine asthe sialogogue, and both antagonist and sialogoguewere injected subcutaneously, in contrast to theintravenous route used by Bulbring et al., and bywhich we have found methantheline to be twiceas effective as atropine.Lehman and Knoeffel's results for the relative

potencies of methantheline and atropine in pre-venting the vasodepressor action of ACh alsodiffer from ours. They found methantheline lesspotent than atropine in this respect, whereas wefound the two drugs to be almost equipotent. Thisdifference is difficult to explain, but may dependon the fact that dogs were used in their experi-ments and cats in ours.From the present work one can say that prop-

antheline is slightly more active than methanthel-ine, but this increase in activity is roughly equallydistributed between its peripheral and autonomicganglion blocking activity. Hambourger (1952)found propantheline approximately five timesmore powerful than methantheline against ACh-induced spasm of guinea-pig gut, and approxi-mately twice as active as methantheline in produc-ing paralysis of the superior cervical ganglion. Wefound no very significant difference between theantimuscarinic activity of propantheline andmethantheline, though our results in the nictitatingmembrane experiments do correlate well withHambourger's. No satisfactory explanation canbe offered for the former anomaly.The local anaesthetic property of methantheline

on the rabbit eye, shown by Lehman and Knoeffel(1944), could not be demonstrated on the guinea-pig eye, though human experiments indicate thatboth propantheline and methantheline have somelocal anaesthetic activity when given intra-cutaneously. How specific this is is not verycertain. It may be that methantheline and prop-antheline damage the nerve endings in the skin,for the injected area remains scarred for severalweeks. The skin does not completely regain itsnormal sensitivity for some time, anaesthesia beingfollowed, for several days after the injection, bya period of hypersensitivity of the protopathictype.The curare-like activity of both compounds is

equal, but very weak (Fig. 6), 2 mg. of each drug

222

METHANTHELINE AND PROPANTHELINE 223

intra-arterially being needed to produce amoderate effect.One main difference between the two drugs

appears to be their duration of action. In themouse mydriasis experiments the duration ofaction of propantheline closely approximated thatof atropine, whereas the action of methanthelinewas much shorter.Although propantheline and, to a lesser extent,

methantheline, approaches hexamethonium inganglion blocking activity, this activity is onlypresent in a dose range, or concentration range,greatly in excess of that necessary for parasym-pathetic blockade. It seems, therefore, that gan-glion blockade is not likely to be responsiblefor any therapeutic effects when these substancesare used clinically-unless a dose range is usedwhich would produce pronounced signs of peri-pheral parasympathetic blockade.

Clinical reports have stressed the advantages ofmethantheline and propantheline as atropine sub-stitutes-in respect of relative absence of side-effects-in doses adequate for treating certaingastrointestinal disorders. Propantheline isclaimed to be better than methantheline from thispoint of view. It may be argued that the relativeinfrequency of serious side-effects with these drugsis further evidence that their ganglionic blockingactivity contributes little if anything to their effec-tiveness in clinical use.

SUMMARY

1. Propantheline and methantheline, two atro-pine-like agents claimed to have in addition con-siderable ganglion blocking activity, have beencompared with atropine and hexamethonium intheir power to produce peripheral parasympatheticblockade and autonomic ganglion blockade respec-tively.

2. Both are more potent antimuscarinic agentsthan atropine, propantheline having 1t to 2 timesthe activity of methantheline.

3. Their ganglion blocking potency, which isexhibited in a much higher dosage range, variesfrom 1/10 to twice that of hexamethonium,depending on the test preparation used. Prop-antheline is, again, the more active.

4. Both drugs have some weak curare-like action.They possess little, if any, local anaestheticactivity.

5. The significance of these findings, in relationto previous work and to the clinical effects ofthe drugs, is discussed.

The authors wish to thank Dr. W. E. Hambourgerand Messrs. G. D. Searle Ltd. for kindly supplyingthe methantheline bromide and propantheline bromideused in these experiments. One of the authors(E.A.J.) carried out this work during the tenure of aMedical Research Council grant for training inresearch methods.

REFERENCES

Bfilbring, E., and Dawes, G. S. (1945). J. Pharmacol.,84, 177.

Burtner, R. R., and Cusic, J. W. (1943). J. Amer.chem. Soc., 65, 1582.

Chance, M. R. A., and Lobstein, H. (1944). J. Pharma-col., 82, 203.

Chittum, J. R., Longino, F. H., Metcalf, B. H., andGrimson, K. S. (1949). Fed. Proc., 8, 25.

Emmens, C. W. (1948). The Principles of BiologicalAssay. London: Chapman and Hall.

Hambourger, W. E. (1952). Personal communication.Cook, D. L., Winbury, M. M., and Freese, H. B.(1950). J. Pharmacol., 99, 245.

Ing, H. R., Dawes, G. S., and Wajda, I. (1945). Ibid.,85, 85.

Lehman, G., and Knoeffel, P. K. (1944). Ibid., 80, 335.Longino, F. H., Grimson, K. S., Chittum, J. R., and

Metcalf, B. H. (1950). Gastroenterol., 14, 301.Lyons, C. K., Reeves, R. J., and Grimson, K. S. (1950).

J. Amer. med. Ass., 143, 873.