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Antagonistic effects of reserpine on d-tubocurarine action on motor function of rabbits
EUROPEAN JOURNAL OF PHARMACOLOGY 6 (1969) 125-128. NORTH-HOLLAND PUBLISHING COMP., AMSTERDAM
ANTAGONISTIC
EFFECTS OF RESERPINE ON MOTOR FUNCTION
ON d-TUBOCURARINE OF RABBITS*
ACTION
Gerald O.CARRIER**, Barbara L.PEGRAM** and Oliver CARRIER Jr.** Department o f Pharmacology, The University of Mississippi Medical Center, Jackson, MississippL and the University of Texas Medical School at San A ntonio, in San Antonio, Texas
Accepted 8 January 1969
Received 25 November 1968
G.O.CARRIER, B.L.PEGRAM and O.CARRIER Jr., Antagonistic effects of reserpine on d-tubocurarine action on motor function of rabbits, European J. Pharmacol. 6 (1969) 125-128. This study was designed to investigate the possibility that reserpine might exhibit an action at the neuromuscular junction by observing its interaction with d-tubocurarine. It was found that the dose of d-tubocuradne to cause paralysis of the hind limbs in normal rabbits was 0.176 + 0.008 mg/kg. However, after the animals received various doses of reserpine, the required dose of d-tubocurarine needed increased significantly. Since reserpine does not alter neuromuscular transmission, it is possible that reserpine increases the excitability of the end plate. Thus, a larger dose of d-tubocurarine is needed for hind limb paralysis. Reserpine
d-Tubocurarme
1. INTRODUCTION Of all the Rauwolfia alkaloids isolated so far, the one which has been most extensively studied both in the laboratory and in the clinic is reserpine. Reserpine is the first alkaloid to display the pecuhar central and hypotensive effects associated with Rauwolfia preparations and later found to be properties of deserpidine and rescinnamine as well (Bein, 1956). Reserpine exhibits a complex pattern of activity. Characteristic features are: a sedative and hypnotic effect, a lowering of arterial blood pressure accompanied by bradycardia, some respiratory inhibition, stimulation of peristalsis, miosis, relaxation of nictitating membrane, and an effect on the temperature regulation center (Bein, 1953). In conducting experiments to further investigate * This work was supported m part by USPHS Grant HE09391. ** Present address: The University of Texas Medical School at San Antomo, Department of Pharmacology, 7703 Floyd Curl Drwe, San Antonio, Texas 78229.
Neuromuscular Junction
various aspects of reserpine's action in our laboratory, we have previously noticed that the animals pretreated with reserpine showed signs of muscle rigidity. This phenomenon suggested that reserpine might exhibit an action at the neuromuscular junction, either directly or indirectly. A review of the literature indicates that reserpine does not alter neuromuscular transmission, and the stores or the amount of acetylcholine released are in no way affected (Bein, 1956; Ledda and Baldi, 1965; Bianchi and Beani, 1966). However, on several occasions it has been noted by a few investigators that reserpine causes a Parkinsonianlike rigidity (Moser and Goldman, 1967; Page, 1966; Thienes and Haley, 1964). This indicates that reserpine affects the homeostasis of skeletal muscle in some manner, possibly by interfering with the calcium equilibrium or by altering the enzyme activity of specific cholinesterase at the neuromuscular junction. It is well known that reserpine causes supersensitivity in smooth muscle (vascular, splenic, nictitating membrane) to catecholamines (Burn and Rand, 1958) and other stimuli (Fleming and Trendelenburg, 1961). Carrier and Shibata (1967) have concluded that the
126
G.O.CARRIER, B.L.PEGRAMand O.CARRIER Jr.
development of supersensitivity by reserpine results from its ability to deplete vascular tissue calcium. Recently, it has been found that reserpine potentiates the action of sodium citrate in causing skeletal muscle tetany (Carrier et al., 1968). Therefore, it is not unreasonable to consider the possibility that reserpine causes supersensitivity in the area of the neuromuscular junction. The following experiment was designed to investigate the effect of reserpine at the neuromuscular junction by observing its interaction with the known neuromuscular blocking agent dtubocurarine.
2. METHODS In the first part of this study, 60 albino rabbits of either sex, weighing between 1.0 and 1.5 kg were used. The experimental animals were divided into 5 groups. The divisions were based either on the dose of reserpine admmistered or the length of administration of reserpine. After the administration of reserpine, d-tubocurarine was given intravenously at a concentration of 0.2 mg/ml until the animals were unable to remain standmg on their hind legs. The dose of d-tubocurarine which caused inhibition of the hind limbs was recorded and the time of recovery from paralysis (return of hind hmb to muscle function) was noted. 15 control rabbits were also given d-tubocurarine until hind limb muscle function was lost. The experimental groups were as follows: Group 1 (4 animals). Received 1.0 mg/kg of reserpine 3 hr prior to the admimstration ofd-tubocurarme. Group 2 (3 animals). Received 1.0 mg/kg of reserpine 24 hr prior to the administration of d-tubocurarine. Group 3 (13 animals). Received 0.1 mg/kg/day of reserpine for 7 days before the administration of dtubocurarme. Group 4 (12 animals). Received 0.1 mg/kg/day of reserpine for 14 days before the administration of d-tubocurarine. Group 5 (13 animals). Received 0.3 mg/kg/day of reserpine for 7 days before the administration of dtubocurarine. Group 6 (15 animals). Control group. Received no reserpine. 3 of these animals were given d-tubocurarine each time an experimental group was used. All
animals were purchased at one time from one supplier. This insured against variable introduction by differences in time or nutrition between control and experimental animals. In addition, to eliminate human bias as much as possible, control animals were mixed with the respective experimental groups prior to testing. Separate observations were recorded by two observers. If a significant discrepancy occurred between observations, the animal tested was disregarded. This only occurred twice. The second part of this study was to determine the lethal dose of d-tubocurarine after the administration of reserpine. 23 albino rabbits of either sex were used. They weighed between 1.0 to 1.5 kg. 6 control animals were given d-tubocurarine until death occurred. The experimental animals were divided into 2 groups. One group (8 animals) received 3 mg/kg lntraperitoneally of reserpine 48 hr prior to the administration of d-tubocurarme. The second experimental group (9 animals) received 0.3 mg/kg/ day of reserpine for 7 days prior to the administration of the d-tubocurarine, d-Tubocurarine was administered to all animals intravenous slowly at a concentration of 0.1 mg/ml until death occurred by loss of heart beat
3. RESULTS In table 1, the effect of reserpine on the action of d-tubocurarine at the nueromuscular junction is shown. The dose of d-tubocurarine required to cause paralysis of the hind limbs in normal rabbits, was found to be 0.176 + 0.008 mg/kg. However, after the animals received reserpine, the required dose of d-tubocurarine rose significantly (see table 1). After the administration of 1.0 mg/kg of reserpine for 3 hr or 24 hr, the amount of d-tubocurarine needed for paralysis of the hind limbs increased, statistically this change was not significant, due to the small number of animals tested. On the other hand, the rabbits treated with 0.1 mg/kg and 0.3 mg/kg over a period of one to two weeks (see table 1) required significantly larger doses of d-tubocurarine to cause paralysis of the hind limbs (p < 0.001-p < 0.01). Under all conditions employed, the average dose of d-tubocurarine required for paralysis of the hind
RESERPINE AND CURARE Table 1 Loss of use of hind limbs with d-tuboeurarine after intraperitoneal administration of reserpine. Group
na
Dose of d-tubocurarine (mg/kg)
Controls
13
0.176 + 0.008 b
p-valuec
Reserpine (1.0 mg/kg) 3 hr
4
0.183 + 0.01
N.S.
Reserpine (1.0 mg/kg) 24 hr
3
0.188 +0.008
N.S.
Reserpine (0.1 mg/kg/day) 7 days
13
0.221 -+0.008
~0.001
Reserpine (0.1 mg/kg/day) 14 days
12
0.204 -+0.007
<0.001
Reserpine (0.3 mg/kg/day) 7 days
13
0.228 -+0.017
<0.01
a Number of animals. b Mean value with standard error of the mean. c Significance of difference from control.
limbs was 0.204 mg/kg after the animals were pretreated with reserpine. Table 2 illustrates the results obtained in determining the lethal dose of d-tubocurarine after the rabbits were pretreated with reserpine. A dose of 0.294 -+ 0.01 mg/kg of d-tubocurarine was lethal for the control animals. 48 hr after the administration of 3 mg/kg reserpine the rabbits required 0.448 -+ 0.02 mg/kg to cause death. Administration of 0.3 mg/kg] day for 7 days increased the lethal dose of d-tubocurarine to 0.409 -+0.03 mg/kg.
4. DISCUSSION The data obtained in the present study indicate that reserpine has some activity either on or affecting skeletal muscle. After treatment of rabbits, either chronically or acutely with reserpine, a greater amount of d-tubocurarine was required to produce the same degree o f muscle paralysis. This was true for both loss of the use of hind limbs and death due to d-tubocurarine.
127
Table 2 Lethal dose of curare after lntraperitoneal administration of reserpine. Group
na
Dose of d-tubocurarine (mg/k~)
p-value c
Controls
6
0.294 + 0.01 b
Reserpine (3 mg/kg) 48 hr
8
0.448 -+0.02
<0.001
Reserpine (0.3 mg/kg/day) 7 days
9
0.409 -+0.03
<0.01
a Number of antmals. b Mean value with standard error of the mean. c Significance of difference from control.
It seems likely, therefore, that pretreatment with reserpine resulted in some change occurring at the skeletal muscle neuromuscular junction of the animals tested which made curare less effective. This could be an electrolyte change, anatomical change, or an enzyme activity change. Carrier and Shibata (1967) have shown that reserpine causes a loss of blood vessel tissue calcium content. They have postulated that this electrolyte depletion results in an increase in membrane permeability with a resultant increase in sensitivity of the tissue to various stimuli. This may well also be the case in skeletal muscle. That is, after a calcium loss from the muscle or its environment, it becomes more permeable and as a result more sensitive to acetylcholine. Thus, it would require a greater amount of curare to prevent the actions of acetylcholine. Carrier et al. (1968) have also found that reserpine has the ability to potentiate the action of sodium citrate to cause tetany in mice. It is well known that this tetany results from a reduction in serum calcium and probably a reduction in the extracellular calcium m the area o f the neuromuscular junction (Kuffler, 1945). It is suggested that reserpine can thus alter the activity of skeletal muscle by affecting calcium homeostasis. After the administration o f reserpine neuromuscular transmission is not altered. The nerve is able to function normally, and acetylcholine is released from the terminal endings and binds to its receptor site on the muscle. Ledda and Baldi (1965) confirmed the
128
G.O CARRIER, B L.PEGRAM and O.CARRIER Jr
statement made by Bein (1956) that reserpine does not modify mammalian neuromuscular transmission. It has been reported that reserpine does not change the acetytcholine stores at the neuromuscular junction (Blanchl and Beani, 1966). Ledda and Baldl concluded that there is no change m the amount of acetylchohne released from the nerve terminal. In view of these earlier studies and the present results, it ~s not unreasonable to suggest that the reserpine is not altering neuromuscular transmission but is altering the excitability of the end plate. In other words, after pretreatment with reserpine the threshold for sp/ke generation is lower at the end plate. This in effect makes acetylchohne more effective m causing depolarization of the skeletal muscle end plate. A greater concentration of d-tubocurarlne is therefore required to depress the activity o f the muscle. Kuffler (•944) concluded that the acetylcholine concentration required to set up &scharges at the neuromuscular junction is reduced after a reduction in ionized calcium. Since reserpine has the ability to deplete calcium at least in vascular muscle, and it has been reported that the concentration of acetylchohne is unaltered (Ledda and Baldi, 1965) by reserpine, one can see how this increase in sensitivity can occur.
ACKNOWLEDGEMENT We would like to acknowledge Dr. A.J.Plummer, CIBA Pharmaceutical Co., Summit, New Jersey, for the generous supply of reserpine.
REFERENCES Beln, H.J, 1956, The pharmacology of Rauwolfia, Pharm. Rev. 8, 435. Bein, H J., 1953, Zur Pharmakologaa des Reserpm, ein neues Alkaloid, aus Rauwolfia serpentma Benth., Expenenta 9, 107. Bianchi, C. and L Beam, 1966, Reserpine and the neuromuscularIunctlon, J. Pharm Pharmacol. 18, 757. Burn, J.H. and M.J.Rand, 1958, The action of sympathomimetic amines in animals treated with reserpine, J. Phys1ol. 144, 314. Carrier, O. and S.Shibata, 1967, A possible role for tissue calcmm m reserpine supersensitivity, J Pharmacol. Exptl Therap. 155, 42 Carner, O., G O.Carrier and A.S.Hume, 1968, Increased toxicity of calcium and c~trate after reserpme administration, Texas Reports Biol. and Med (m press). Fleming, W H. and U.Trendelenburg, 1961, The development of supersensitivity to norepmephrine after pretreatment with reserpine, J Pharmacol. Exptl. Therap, 133, 41 Kuffler, S.W., 1944, The effect calcium has on the neuromuscular junction, J. Neurophysiol 7, 17 Kuffler, S.W., 1945, Excitability changes at the neuromuscular Junction durang tetany, J. Physiol. 103,403 Ledda, F. and V.Baldl, 1965, The effects of reserpine on mammalian neuromuscular transmission, Life Sci. 4,955 Moser, M. and A.G.Goldman, 1967, Hypertensive vascular disease, (J.B.Llppincott Co., Philadelphia), pp 194-195 Page, I.H, 1966, Drug treatment of hypertension in antihypersensztlve therapy, ed. F.Gross (Springer-Verlag, New York), pp. 602-614. Thlenes, C.H. and T.J.Haley, 1964, Clinical toxicology (Lea and Feblger, Philadelphia), pp. 109-111.
ANTAGONISTIC
EFFECTS OF RESERPINE ON MOTOR FUNCTION
ON d-TUBOCURARINE OF RABBITS*
ACTION
Gerald O.CARRIER**, Barbara L.PEGRAM** and Oliver CARRIER Jr.** Department o f Pharmacology, The University of Mississippi Medical Center, Jackson, MississippL and the University of Texas Medical School at San A ntonio, in San Antonio, Texas
Accepted 8 January 1969
Received 25 November 1968
G.O.CARRIER, B.L.PEGRAM and O.CARRIER Jr., Antagonistic effects of reserpine on d-tubocurarine action on motor function of rabbits, European J. Pharmacol. 6 (1969) 125-128. This study was designed to investigate the possibility that reserpine might exhibit an action at the neuromuscular junction by observing its interaction with d-tubocurarine. It was found that the dose of d-tubocuradne to cause paralysis of the hind limbs in normal rabbits was 0.176 + 0.008 mg/kg. However, after the animals received various doses of reserpine, the required dose of d-tubocurarine needed increased significantly. Since reserpine does not alter neuromuscular transmission, it is possible that reserpine increases the excitability of the end plate. Thus, a larger dose of d-tubocurarine is needed for hind limb paralysis. Reserpine
d-Tubocurarme
1. INTRODUCTION Of all the Rauwolfia alkaloids isolated so far, the one which has been most extensively studied both in the laboratory and in the clinic is reserpine. Reserpine is the first alkaloid to display the pecuhar central and hypotensive effects associated with Rauwolfia preparations and later found to be properties of deserpidine and rescinnamine as well (Bein, 1956). Reserpine exhibits a complex pattern of activity. Characteristic features are: a sedative and hypnotic effect, a lowering of arterial blood pressure accompanied by bradycardia, some respiratory inhibition, stimulation of peristalsis, miosis, relaxation of nictitating membrane, and an effect on the temperature regulation center (Bein, 1953). In conducting experiments to further investigate * This work was supported m part by USPHS Grant HE09391. ** Present address: The University of Texas Medical School at San Antomo, Department of Pharmacology, 7703 Floyd Curl Drwe, San Antonio, Texas 78229.
Neuromuscular Junction
various aspects of reserpine's action in our laboratory, we have previously noticed that the animals pretreated with reserpine showed signs of muscle rigidity. This phenomenon suggested that reserpine might exhibit an action at the neuromuscular junction, either directly or indirectly. A review of the literature indicates that reserpine does not alter neuromuscular transmission, and the stores or the amount of acetylcholine released are in no way affected (Bein, 1956; Ledda and Baldi, 1965; Bianchi and Beani, 1966). However, on several occasions it has been noted by a few investigators that reserpine causes a Parkinsonianlike rigidity (Moser and Goldman, 1967; Page, 1966; Thienes and Haley, 1964). This indicates that reserpine affects the homeostasis of skeletal muscle in some manner, possibly by interfering with the calcium equilibrium or by altering the enzyme activity of specific cholinesterase at the neuromuscular junction. It is well known that reserpine causes supersensitivity in smooth muscle (vascular, splenic, nictitating membrane) to catecholamines (Burn and Rand, 1958) and other stimuli (Fleming and Trendelenburg, 1961). Carrier and Shibata (1967) have concluded that the
126
G.O.CARRIER, B.L.PEGRAMand O.CARRIER Jr.
development of supersensitivity by reserpine results from its ability to deplete vascular tissue calcium. Recently, it has been found that reserpine potentiates the action of sodium citrate in causing skeletal muscle tetany (Carrier et al., 1968). Therefore, it is not unreasonable to consider the possibility that reserpine causes supersensitivity in the area of the neuromuscular junction. The following experiment was designed to investigate the effect of reserpine at the neuromuscular junction by observing its interaction with the known neuromuscular blocking agent dtubocurarine.
2. METHODS In the first part of this study, 60 albino rabbits of either sex, weighing between 1.0 and 1.5 kg were used. The experimental animals were divided into 5 groups. The divisions were based either on the dose of reserpine admmistered or the length of administration of reserpine. After the administration of reserpine, d-tubocurarine was given intravenously at a concentration of 0.2 mg/ml until the animals were unable to remain standmg on their hind legs. The dose of d-tubocurarine which caused inhibition of the hind limbs was recorded and the time of recovery from paralysis (return of hind hmb to muscle function) was noted. 15 control rabbits were also given d-tubocurarine until hind limb muscle function was lost. The experimental groups were as follows: Group 1 (4 animals). Received 1.0 mg/kg of reserpine 3 hr prior to the admimstration ofd-tubocurarme. Group 2 (3 animals). Received 1.0 mg/kg of reserpine 24 hr prior to the administration of d-tubocurarine. Group 3 (13 animals). Received 0.1 mg/kg/day of reserpine for 7 days before the administration of dtubocurarme. Group 4 (12 animals). Received 0.1 mg/kg/day of reserpine for 14 days before the administration of d-tubocurarine. Group 5 (13 animals). Received 0.3 mg/kg/day of reserpine for 7 days before the administration of dtubocurarine. Group 6 (15 animals). Control group. Received no reserpine. 3 of these animals were given d-tubocurarine each time an experimental group was used. All
animals were purchased at one time from one supplier. This insured against variable introduction by differences in time or nutrition between control and experimental animals. In addition, to eliminate human bias as much as possible, control animals were mixed with the respective experimental groups prior to testing. Separate observations were recorded by two observers. If a significant discrepancy occurred between observations, the animal tested was disregarded. This only occurred twice. The second part of this study was to determine the lethal dose of d-tubocurarine after the administration of reserpine. 23 albino rabbits of either sex were used. They weighed between 1.0 to 1.5 kg. 6 control animals were given d-tubocurarine until death occurred. The experimental animals were divided into 2 groups. One group (8 animals) received 3 mg/kg lntraperitoneally of reserpine 48 hr prior to the administration of d-tubocurarme. The second experimental group (9 animals) received 0.3 mg/kg/ day of reserpine for 7 days prior to the administration of the d-tubocurarine, d-Tubocurarine was administered to all animals intravenous slowly at a concentration of 0.1 mg/ml until death occurred by loss of heart beat
3. RESULTS In table 1, the effect of reserpine on the action of d-tubocurarine at the nueromuscular junction is shown. The dose of d-tubocurarine required to cause paralysis of the hind limbs in normal rabbits, was found to be 0.176 + 0.008 mg/kg. However, after the animals received reserpine, the required dose of d-tubocurarine rose significantly (see table 1). After the administration of 1.0 mg/kg of reserpine for 3 hr or 24 hr, the amount of d-tubocurarine needed for paralysis of the hind limbs increased, statistically this change was not significant, due to the small number of animals tested. On the other hand, the rabbits treated with 0.1 mg/kg and 0.3 mg/kg over a period of one to two weeks (see table 1) required significantly larger doses of d-tubocurarine to cause paralysis of the hind limbs (p < 0.001-p < 0.01). Under all conditions employed, the average dose of d-tubocurarine required for paralysis of the hind
RESERPINE AND CURARE Table 1 Loss of use of hind limbs with d-tuboeurarine after intraperitoneal administration of reserpine. Group
na
Dose of d-tubocurarine (mg/kg)
Controls
13
0.176 + 0.008 b
p-valuec
Reserpine (1.0 mg/kg) 3 hr
4
0.183 + 0.01
N.S.
Reserpine (1.0 mg/kg) 24 hr
3
0.188 +0.008
N.S.
Reserpine (0.1 mg/kg/day) 7 days
13
0.221 -+0.008
~0.001
Reserpine (0.1 mg/kg/day) 14 days
12
0.204 -+0.007
<0.001
Reserpine (0.3 mg/kg/day) 7 days
13
0.228 -+0.017
<0.01
a Number of animals. b Mean value with standard error of the mean. c Significance of difference from control.
limbs was 0.204 mg/kg after the animals were pretreated with reserpine. Table 2 illustrates the results obtained in determining the lethal dose of d-tubocurarine after the rabbits were pretreated with reserpine. A dose of 0.294 -+ 0.01 mg/kg of d-tubocurarine was lethal for the control animals. 48 hr after the administration of 3 mg/kg reserpine the rabbits required 0.448 -+ 0.02 mg/kg to cause death. Administration of 0.3 mg/kg] day for 7 days increased the lethal dose of d-tubocurarine to 0.409 -+0.03 mg/kg.
4. DISCUSSION The data obtained in the present study indicate that reserpine has some activity either on or affecting skeletal muscle. After treatment of rabbits, either chronically or acutely with reserpine, a greater amount of d-tubocurarine was required to produce the same degree o f muscle paralysis. This was true for both loss of the use of hind limbs and death due to d-tubocurarine.
127
Table 2 Lethal dose of curare after lntraperitoneal administration of reserpine. Group
na
Dose of d-tubocurarine (mg/k~)
p-value c
Controls
6
0.294 + 0.01 b
Reserpine (3 mg/kg) 48 hr
8
0.448 -+0.02
<0.001
Reserpine (0.3 mg/kg/day) 7 days
9
0.409 -+0.03
<0.01
a Number of antmals. b Mean value with standard error of the mean. c Significance of difference from control.
It seems likely, therefore, that pretreatment with reserpine resulted in some change occurring at the skeletal muscle neuromuscular junction of the animals tested which made curare less effective. This could be an electrolyte change, anatomical change, or an enzyme activity change. Carrier and Shibata (1967) have shown that reserpine causes a loss of blood vessel tissue calcium content. They have postulated that this electrolyte depletion results in an increase in membrane permeability with a resultant increase in sensitivity of the tissue to various stimuli. This may well also be the case in skeletal muscle. That is, after a calcium loss from the muscle or its environment, it becomes more permeable and as a result more sensitive to acetylcholine. Thus, it would require a greater amount of curare to prevent the actions of acetylcholine. Carrier et al. (1968) have also found that reserpine has the ability to potentiate the action of sodium citrate to cause tetany in mice. It is well known that this tetany results from a reduction in serum calcium and probably a reduction in the extracellular calcium m the area o f the neuromuscular junction (Kuffler, 1945). It is suggested that reserpine can thus alter the activity of skeletal muscle by affecting calcium homeostasis. After the administration o f reserpine neuromuscular transmission is not altered. The nerve is able to function normally, and acetylcholine is released from the terminal endings and binds to its receptor site on the muscle. Ledda and Baldi (1965) confirmed the
128
G.O CARRIER, B L.PEGRAM and O.CARRIER Jr
statement made by Bein (1956) that reserpine does not modify mammalian neuromuscular transmission. It has been reported that reserpine does not change the acetytcholine stores at the neuromuscular junction (Blanchl and Beani, 1966). Ledda and Baldl concluded that there is no change m the amount of acetylchohne released from the nerve terminal. In view of these earlier studies and the present results, it ~s not unreasonable to suggest that the reserpine is not altering neuromuscular transmission but is altering the excitability of the end plate. In other words, after pretreatment with reserpine the threshold for sp/ke generation is lower at the end plate. This in effect makes acetylchohne more effective m causing depolarization of the skeletal muscle end plate. A greater concentration of d-tubocurarlne is therefore required to depress the activity o f the muscle. Kuffler (•944) concluded that the acetylcholine concentration required to set up &scharges at the neuromuscular junction is reduced after a reduction in ionized calcium. Since reserpine has the ability to deplete calcium at least in vascular muscle, and it has been reported that the concentration of acetylchohne is unaltered (Ledda and Baldi, 1965) by reserpine, one can see how this increase in sensitivity can occur.
ACKNOWLEDGEMENT We would like to acknowledge Dr. A.J.Plummer, CIBA Pharmaceutical Co., Summit, New Jersey, for the generous supply of reserpine.
REFERENCES Beln, H.J, 1956, The pharmacology of Rauwolfia, Pharm. Rev. 8, 435. Bein, H J., 1953, Zur Pharmakologaa des Reserpm, ein neues Alkaloid, aus Rauwolfia serpentma Benth., Expenenta 9, 107. Bianchi, C. and L Beam, 1966, Reserpine and the neuromuscularIunctlon, J. Pharm Pharmacol. 18, 757. Burn, J.H. and M.J.Rand, 1958, The action of sympathomimetic amines in animals treated with reserpine, J. Phys1ol. 144, 314. Carrier, O. and S.Shibata, 1967, A possible role for tissue calcmm m reserpine supersensitivity, J Pharmacol. Exptl Therap. 155, 42 Carner, O., G O.Carrier and A.S.Hume, 1968, Increased toxicity of calcium and c~trate after reserpme administration, Texas Reports Biol. and Med (m press). Fleming, W H. and U.Trendelenburg, 1961, The development of supersensitivity to norepmephrine after pretreatment with reserpine, J Pharmacol. Exptl. Therap, 133, 41 Kuffler, S.W., 1944, The effect calcium has on the neuromuscular junction, J. Neurophysiol 7, 17 Kuffler, S.W., 1945, Excitability changes at the neuromuscular Junction durang tetany, J. Physiol. 103,403 Ledda, F. and V.Baldl, 1965, The effects of reserpine on mammalian neuromuscular transmission, Life Sci. 4,955 Moser, M. and A.G.Goldman, 1967, Hypertensive vascular disease, (J.B.Llppincott Co., Philadelphia), pp 194-195 Page, I.H, 1966, Drug treatment of hypertension in antihypersensztlve therapy, ed. F.Gross (Springer-Verlag, New York), pp. 602-614. Thlenes, C.H. and T.J.Haley, 1964, Clinical toxicology (Lea and Feblger, Philadelphia), pp. 109-111.