Some Effects of Quinidine and Quinine on the Electromyogram of the Colon

GASTROENTEROLOGY 65: 773-777, 1973 Copyright© 1973 by The Willia ms & Wilkins Co.

Vol. 65, No.5 Printed in U.S.A.

SOME EFFECTS OF QUINIDINE AND QUININE ON THE ELECTROMYOGRAM OF THE COLON JoHN D. BARKER, JR., M.D. , AND JAMES CHRISTENSEN , M.D.

Gastroenterology Research Laboratories, Department of Internal Medicine, University and Veterans Administration Hospitals, University of Iowa, Iowa City, Iowa

The cat colon was exposed in vitro to two diarrheogenic agents, quinidine and quinine. Six electrodes, uniformly spaced in the long axis of the ascending colon, recorded the electromyogram before and after the addition of either quinidine hydrochloride or quinine dihydrochloride. Records were read for changes in electrical slow wave frequency and congruence, and for changes in the proportion of slow waves bearing spike potentials. Both agents significantly raised slow wave frequency at concentrations of 5 x I0- 4 M. Slow wave congruence (as measured by calculating the coefficient of variation of frequency among the six electrodes) was significantly reduced by quinidine at 2 x I0- 4 M and by quinine at 5 x I0- 4 M. The proportion of slow waves bearing spike potentials increased significantly with quinidine at 5 x I0- 4 M but not at I0- 3 M . The proximal cat colon examined in vitro generates electrical slow waves spreading orad from a single pacemaker most of the time. These constantly recurring signals originate in the circular muscle layer. Rhythmic contractions of the circular muscle layer in the proximal colon accompany spike potentials. Spike potentials in this part of the colon occur only during 1 part of the slow wave cycle. 2• 4 Thus, the distribution of slow waves in time and space should influence the distribution of contractions, and this distribution may be important in fecal flow . Colons taken from cats with spontaneous diarrhea and examined in vitro show altered spreading patterns of slow waves .5 A Received March 5, 1973. Accepted June 27, 1973. Presented in part in abstract form .' Address requests for reprints to: Dr. James Christensen , Department of Internal Medicine, University Hospitals, Iowa City, Iowa 52242. This work was supported in part by Veterans Administration research funds, in part by a Veterans Administration Training Grant in Gastroenterology, and in part by National Institute of Health Research Career Development Award AM-20547 . 773

similar alteration occurs in colons taken from cats given castor oil by mouth and examined in vitro. 5 The sodium salt of hydroxyoleic acid, the active principal of castor oil, induces the same change in the cat colon in vitro .6 Thus, diarrhea may, in part, be a consequence of a change in spreading patterns of electrical slow waves in the proximal colon . Since quinidine and quinine induce diarrhea, we examined their effects on spreading patterns of electrical slow waves in the proximal colon.

Methods Healthy cats, 1.2 to 5.5 kg, were anesthetized by intrapleural injection of sodium pentobarbi· tal , 30 mg per kg. The colon was measured and marked in situ and removed by transection of the ileum 1 em above the ileocecal junction and of the distal colon at the pelvic brim. Colon function was judged by the consistency of the stool; colons containing liquid stool were discarded. The cecal tip was amputated and the colon was everted over a hollow and perforated plexiglass mandrel. Colons of various sizes could be accommodated by insertion of the tapered mandrel to an appropriate distance .

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The colon was brought to its approximate dimensions in situ and immersed in a bath of Krebs solution, aerated with 95% 0 2 - 5% CO. at 36 to 38 C. Stool remaining on the mucosa was removed with a sponge. A 1.5- by 6.0-cm patch of mucosa was removed to expose the submucosal vessels lying on the circular muscle layer. The long axis of this exposed surface lay in the long axis of the colon over the mesenteric insertion , beginning about 0.5 em below the ileocecal junction. The prepared colon was transferred to a jacketed bath of Krebs solution, aerated with 95 % 0. - 5% CO. at 36.5 to 37.5 C. Six silver chloride glass pore electrodes•· • were aligned vertically 8 mm apart over the mesenteric insertion with the first electrode about 6 mm below the ileocecal junction. Each electrode was lowered until slow waves were clearly detected. Electrical potential difference was recorded separately between each of the six electrodes and a common grounded reference electrode, a coil of chlorided silver wire, was immersed in the bath. The electrodes were connected to the RC amplifiers of an ink-writing polygraph . The time constant was 1 sec and high frequency filters dampened transients faster than about 30 Hz. The time from removal of the colon to the beginning of the recording was 15 min or less. Thirty minutes were allowed for stabilization of each colon after recording began. A subsequent 60-min control pe~iod was followed by a 60-min treatment period . Aliquots of fresh stock solutions of quinidine or quinine were added to the bath at the end of the control period. Each colon received only one agent at one concentration . There were five experiments with no drug added in the treatment period. There were five experiments with each of the following concentrations of quinidine hydrochloride (as the salt): 10- s M, 5 X 10-s M, 10- • M , 2 X 10- • M , 5 X 10- • M, and 10- • M . There were another four studies with no drug added in the treatment period and four others with each of the following concentrations of quinine dihydrochloride (as the salt): w-s M, lQ- • M, and 5 X w-· M . Volumes of drug solutions added to the 1.2-liter bath never exceeded 60 mi. In . order to achieve a bath COncentration of 10-a M quinidine hydrochloride, the entire bath had to be exchanged because of difficulty in solubilization. In the exchange, bath temperature did not change more than 1 C and the time required was kept below 30 sec. Slow wave frequency was measured by counting the slow waves recorded by each of the six electrodes separately in sequential 5-min periods through both control and treatment hours.

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Then the mean frequency in all5-min periods in each hour was calculated separately for control and treatment periods at each electrode. These values were averaged among all electrodes separately in the control and treatment periods. The difference in mean frequency between control and treatment periods was expressed as percentage of change from the control period: Spike burst activity was measured by computing the proportion of all slow waves bearing spike potentials in each sequential 5-min period. Mean values were calculated as percentage of all slow waves for the control and treatment periods separately. Since spike bursts signal contractions, the incidence of spike bursts reflects contractile activity. Records were also read for congruence of slow waves . The terms "congruence" and "complete coupling" were used previously to describe the state of phase-lock of slow waves in similar studies in this part of the colon. •-• Briefly stated, congruence exists when slow waves are phase-locked across all six recording sites, appearing to originate from a single source. Whim slow waves at all six points appear to arise from two or more sources , the slow waves are said to be incongruent or incompletely coupled. The method used previously to examine this property of slow waves yielded only an estimate Of the proportion of time when slow waves were congruent. The technique used in these experiments depends upon the fact that when there are several sources for slow waves the sources generate slow waves at different frequencies . To discover incongruence , we calculated the coefficient of variation of frequency among all six electrodes in 5-min periods throughout the control and treatment periods. The coefficient of variation was determined by calculating the average of the frequencies at each recording point separately for each 5-min period throughout control and treatment periods. The average frequency among the six electrodes in each 5-min period was divided into the standard deviation and the quotient was multiplied by 100. The mean coefficients of variation for all 5-min intervals in the control and treatment periods were computed and compared. When frequency is identical at all six points, the coefficient of variation is zero and congruence exists. As the number of sources for slow waves increases, frequency among the six electrodes varies, the coefficient of variation of frequency at the six points increases and incongruence exists. Significance of differences for slow wave frequency, incidence of spike bursts, and slow

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QUINIDINE, QUININE, AND THE COLON

wave congruence was determined by the Wil - bearing spike potentials also increased coxon ranking method. 7 after exposure to quinidine. This change

Results Quinidine. The addition of quinidine hydrochloride to the bath produced marked changes in the electromyogram (fig. 1) . This effect was evident as early as 3 min after the addition. Quinidine raised the frequency of slow waves. This change was apparent at a concentration of 10- 4 M in some experiments and, considering all experiments, achieved a level of significance (P < 0.05) at 5 x 10- 4 M and above (fig. 2). The proportion of slow waves 35

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also was apparent in some experiments at 10- 4 M, and, considering all experiments, was significant (P < 0.01) at 5 x 10- 4 M but not at 1 x 10- a M. Quinidine also increased the coefficient of variation of frequency among the six recording points. This change was apparent in some experiments at 10- 4 M, and, considering all experiments, reached a level of significance (P < 0.05) at 2 x 10- 4 M and above (fig. 3). Quinine. Quinine produced changes in the electromyogram like those seen with quinidine. The effect was evident within 3 34

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FIG . 1. A record of the colon electromyogram before and after exposure of the colon to quinidine hydrochloride, 5 x 10· • M . A is the record from 32 to 36 min before, and B is the record from 32 to 36 min after the addition of quinidine hydrochloride to make a bath concentration of 5 x 10· • M. The numbers 1 through 6 at the left of each record represent the numbers of the electrodes: electrode 1 is 6 mm below the ileocecal junction , and the electrodes are 8 mm apart. The voltage calibration is the same for all electrodes. The time scale at the top shows seconds and numbered minutes. Dashed lines have been drawn across panel A to show the phase-lock of slow waves to a source distal to the region spanned by the electrodes . In panel B, three sources seem to be operating, one driving the slow waves at electrode 1, one governing those at electrode 2, and a third controlling those at electrodes 3 through 6.

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Vol. 65, No.5

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FIG . 2. The effect of quinidine hydrochloride on slow wave frequency. The horizontal axis shows the molar concentration of the agent on a log scale . The vertical axis shows the ratio of the mean frequency in the treatment period to that in the control period, expressed as percentage of change. Points on the graph indicate mean data from each of five experiments done at each concentration indicated, including five control experiments. Lines connect median points. Asterisks indicate data significantly different from control data (P < 0.05) .

FIG . 3. The effect of quinidine hydrochloride on congruence of slow waves. The horizontal axis indicates molar concentrations of quinidine hydrochloride on a log scale. The vertical axis shows the ratio (treatment period-control period) of the coefficient of variation of frequency among the six electrodes. The ratio is expressed as percentage of change from control to treatment periods. Points represent data for each of five experiments at each concentration and five control experiments. The line connects median points. Asterisks indicate data significantly different from control data (P < 0.05) .

tended to increase but the change did not reach a level of significance at the highest concentration used, 5 x 10- 4 M. The coefficient of variation of frequency among the electrodes was increased by quinine and reached a level of significance (P < 0.01) at 5 X 10- 4 M (fig. 4).

Discussion Both quinidine and quinine increase the coefficient of variation of slow wave frequency along the proximal colon . Since phase-lock of slow waves would require that frequency be the same in this segment, this effect represents a reduced state of phase-lock or incongruence of slow waves. This is the change seen previously in animals with spontaneous diarrhea and in those treated with castor oil, and is the same change induced by the exposure of the colon in vitro to hydroxyoleic acid. Both quinidine and quinine tend to increase the mean frequency of the slow waves. Only quinidine increases the incidence of spike bursts. Both quinine and quinidine may cause diarrhea in therapy. Quinine, a stereoisomer of quinidine, had occasional use as a laxative, enema, or suppository in the past. Garrett et a!., 8 using chronically implanted electrodes in vivo, examined the effect of intravenously administered quinidine on slow waves of the dog small intestine. Quinidine did not change the fre• (18951

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QUINIDINE, QUININE, AND THE COLON

quency of slow waves, but it increased the proportion of slow waves bearing spike potentials and it excited contractions of the small bowel. Those authors did not examine phase-lock or congruence of slow waves. They proposed that quinidine increased the responsiveness of the small bowel to slow waves. Quinidine is concentrated in skeletal muscle, liver, kidney, and cardiac muscle. 9 Therapeutic blood levels range from 1.3 to 5.4 mg per 1 and cardiotoxicity occurs above 15 mg per liter. The bath concentrations we used ranged from about 3.9 to 389 mg per liter. The effects we demonstrated occurred at concentrations above 39 mg per liter. Thus, these were induced by concentrations in the cardiotoxic ranges of serum concentrations. The mechanism of the colonic electromyographic effects of the agents is unknown. Quinidine chelates calcium, 10 but a reduction of calcium concentration reduces the frequency of colon slow waves, and slow waves disappear in a calciu.p1-free medium. 11 Thus, it is doubtful that calcium chelation explains the electromyographic changes observed. Both quinidine and quinine have weak anticholinergic actions. This also seems unlikely as an explanation since atropine has little effect on the electromyogram except to counteract the excitatory actions of cholinergic agents. 12 The effect of atropine on slow wave congruence is not known. The principal effect of the diarrheogenic agents investigated appears to be a reduced phase-lock of slow waves in the proximal colon. The emergence of multiple pacemakers diminishes integration of slow waves along the proximal colon. Since slow waves pace contractions, this should lead

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to diminished integration of wall movement. If the integration of wall movement affects fecal flow, loss of integrated wall movement might result in liquid stool. This might contribute to the diarrhea seen in quinidine and quinine therapy. REFERENCES 1. Barker JD Jr, Christensen J: Effects of quinidine on colon slow waves and spike potentials in vitro (abstr). Gastroenterology 62:837, 1972 2. Christensen J, Caprilli R, Lund GF : Electric slow waves in circular muscle of cat colon. Am J Phys iol 217:771-776, 1969 3. Christensen J , Hauser RL: Longitudinal axial coupling of slow waves in proximal cat colon. Am J Physiol 221 :246-250, 1971 4. Christensen J , Hauser RL: Circumferential coupling of electric slow waves in circular muscle of cat colon. Am J Physiol 221: 1033-1037, 1971 5. Christensen J, Weisbrodt NW, Hauser RL: Electric al slow wave of the proximal colon of the cat in diarrhea. Gastroenterology 62:1167-1173, 1972 6. Christensen J , Freeman BW: Circular muscle electromyogram in the cat colon: local effect of sodium ricinoleate. Gastroenterology 63:10111015, 1972 7. Wilcoxon F, Wilcox RA: Some Rapid Approximate Statistical Procedures. Lederle Laboratories, 1964, p 7- 17 8. Garrett JM , Schlegel JF, Code CF: Effect of quinidine on electrical and motor activity of canine small bowel. Gut 7:562-565, 1966 9. Kay CF: Quinidine: a re-evaluation. Med Clin North Am 50:1221-1230, 1966 10. Conn HL, Luchi RJ: Some cellular and metabolic considerations relating to the action of quinidine as a prototype antiarrhythmic agent. Am J Med 37:685-699, 1964 11. Wienbeck M, Christensen J: Cationic requirements of colon slow waves in the cat. Am J Physiol 220:513-519, 1971 12. Wienbeck M, Christensen J: Effects of some drugs on electrical activity of the isolated colon of the cat. Gastroenterology 61:470-478, 1971