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Effects of microiontophoretic application of imipramine on single neurones in the brain stem
Neuropharmacology,
1971,10,661-664Pergamon Press.Printed
in Gt. Britain.
PRELIMINARY NOTE EFFECTS OF MICROIONTOPHORETIC APPLICATION OF IMIPRAMINE ON SINGLE NEURONES IN THE BRAIN STEM G. L. AVANZINO, ROSA ERMIRIOand C. ZUMMO Istitutodi Fisiologia Umana dell’Universitl di Genova, Italy (Accepted 24 February 197 1) Sununary-Imipramine was applied by microiontophoresis to single neurones of the brain stem in guinea pigs under local anaesthesia. The effects obtained were compared to those of noradrenaline applied microiontophoretically to the same neurone. The interaction of these two drugs was also studied. NA and IMI produced similar effects and a synergism between these two drugs was noted. These effects may be due to a blockade of the neurone uptake of NA by IMI. THEREis
considerable evidence to show that imipramine (IMT) antagonizes the uptake of noradrenaline (NA) by the postganglionic sympathetic innervation of various organs (GLOWINSKIand BALDESSARINI, 1966; IVERSEN, 1968). According to the “uptake hypothesis”, the main physiological function of NA uptake is to terminate the action of NA released from presynaptic adrenergic nerve terminals. Results that support this hypothesis have been obtained mainly in the peripheral nervous system (LANGERet al., 1967; IVERSEN,1968). Evidence for NA uptake in the CNS has been obtained by biochemical, histochemical and autoradiographic studies; in this case too, IMI seems to act as a blocker of NA uptake (GLOWINSKIand AXELROD, 1964; GLOWINSKI and BALDESSARINI, 1966; Ross and RENYI, 1967; CARLSS~Net al., 1968, 1969).
The present study concerns the effects of IMI on the impulse frequency of neurones in the CNS when this substance is released by microiontophoresis very close to the neurone under study. The effects of IMI were compared to those of NA applied microiontophoretically to the same neurone. The interaction of the two compounds was also studied. Results were obtained from 10 guinea pigs of either sex. Craniectomy was performed and the cerebellum was sucked out under ether anaesthesia. Wound margins and the pressure points of the head clamp were infiltrated with novocaine and the animal was immobilized with an i.p. injection of sincurarina (Farmitalia, SA Milano). The ether anaesthesia was then discontinued, and the animal was ventilated by a respiratory pump. For microiontophoresis, multibarrelled micropipettes (prepared according to BRADLEY et al., 1966) were used. One barrel of the micropipettes contained IMI hydrochloride, 7-IO%, pH 5, (Geigy, SA Milano); another contained NA bitartrate, IO%, pH 7, (KochLight Laboratories Ltd., Colnbrook, Buckinghamshire); a third barrel contained 1 M NaCl for monitoring current effects, and unitary discharges were recorded through a barrel containing 4 M NaCl. The micropipettes were inserted through the floor of the fourth ventricle. 661
662
G. L. AVANZINO,ROSAERMXRIO and C. ZUMMO
Fifty-one neurones, belonging mainly to the brain stem reticular formation, were studied Microiontophoretic appiication of NA by positive currents between 25 and 80 nA had inhibitory effects on 38 neurones, excitatory effects on 8, and no clear action on 5 neurones. Application of IMI with positive currents between 25 and 70 nA usually produced effects qualitatively similar to those of NA. IMI inhibited 26 of the neurones inhibited by NA and excited 5 of the neurones excited by NA. The effects of IMI were often weaker than those of NA (Figs. lA, B). Simultaneous application of NA and IMI produced longer or greater
A NA
-!!!L
N&I
30 -
f 20-
B
0
x)0
xx)
ma
-t FIG. 1. Effect of microiontophoretic application of noradrenaline (NA) and imipramine (IMI) on the luring frequency of 2 untts in the brain stem of a guinea pig. NA, applied at 40 nA for 30 set, had inhibitory (A) and excitatory (B) effects. IMI, applied at 30 nA for 40 see, weakly mimicked these actions of NA. The application of both substances together caused longer and more marked effects. In (A), current effects were tested by applying an amount of current equal to the total current used in applying NA and IMI. Axes: f, the total number of impulses every 5 set; t, the time in sec.
Imipramineand brain stem neurones
663
inhibition than was produced by the application of either compounds alone in 15 out of 18 neurones tested. Prolonged or increased excitation was seen in 2 neurones out of 6. When applied at 40-50 nA for 30 set, between 20 and 180 see before NA, IMI usually did not increase the NA effect (Fig. 2). When applied at 60-l 00 nA for 80-120 set i~~diately before NA, IMI occasionally depressed the response to NA,
NA
NA
30
f
20
10
~
FIG. 2. Effects of the application of nomdrenaline(NA}at 50 nA for 25 see and i~pr~ne (IMI}at 40nA for 3Osec.A second application of NA i2Osecafter the end of the IMI application had a similare&et to the first applicationof NA.
The current e%xts of simultaneous applications of NA and IMI were checked by passing the appropriate total current through the barrel containing 1 M NaCl (Fig. 1A) or by simultaneous application of current and NA, and of current and IMI. These results suggest that NA and IMI have qualitatively similar effects when applied by microiontophoresis to singIe neurones, and that there is synergism between these effects. It is suggested that these effects could be due to a blockade of the neuronal uptake of NA by IMI. The results so far obt~ned are not su~cient to determine whether the reduction of the effects of NA after prolonged applications of IMX are due to de~nsiti~tion of the NA effect or to an antagonistic action by IMI. REFERENCES BRADLEY,P, B., DHAWAN,B. N. and WOLSTENCROFT, J. IS. (1966). Pharmacological properties of cholinoceptive neurones in the medulla and pons of the cat. J. Physiol., Land. 183: 658-674. CARLWON, A., FUXE,K. and UNOERSTEDT, U. (1968). The effect of Imipramine on central 5 hydroxytryptamine neurons. J. Pharm. Pharmac. 20: 150-l 51. CARLSSON, A,, CORRODI, A., FUXE,K. and HBKFELT,T. (1969). ES?& of some antidepressant drugs on the depletion of intraneuronal brain catecholamine stores caused by 4 p dimethyl-men-tym~ne. Eur. J, Pharntac. 5: 367-373. GLOWIN~KI,
J. and AXELR~D,J. (1964). Inhibition of uptake of tritiated-noradrenaline in the intact rat brain by imipramine and structurafly related compounds. Nature, Load. 204: 1318-1319. ~~WINSKI, 3. and BALDESSARINI, R. J. (1966). Metallic of ~orepinep~ine in the Central Nervous System. Phar~ac. Rev. 18: 1201-1238.
664 fvmm,
G. L. AYANUNO, ROSA ERM~KXOand C. Zu~hno
L. It. (I%tQ. R&z #~~~~~~~~~~~ uptake in adrenergic n~~~srn~ss~~~. In: CTBA F&t. Symp. Adrenergic ~eur~transrni~~~n~ pp. 44-50. Churchill. London. LAwER, S. Z., DRASKOCZ~,R. R. and ~~~EL~NB~~~ U. f1967). Time course of the de~ekjpment of wpersensitivity to various amines in the nictitating membrane of the pithed cat after denervation or decentralization. 3. Phamac, exp. Tlrer, 157: 255-273. RCW, S, B, and RENYI, A. L. (1967). Inhibition ofthe uptake oftritiated catechotamines by antidepressant and related agents. Eur. J. Pharmac. 2: 181--186.
1971,10,661-664Pergamon Press.Printed
in Gt. Britain.
PRELIMINARY NOTE EFFECTS OF MICROIONTOPHORETIC APPLICATION OF IMIPRAMINE ON SINGLE NEURONES IN THE BRAIN STEM G. L. AVANZINO, ROSA ERMIRIOand C. ZUMMO Istitutodi Fisiologia Umana dell’Universitl di Genova, Italy (Accepted 24 February 197 1) Sununary-Imipramine was applied by microiontophoresis to single neurones of the brain stem in guinea pigs under local anaesthesia. The effects obtained were compared to those of noradrenaline applied microiontophoretically to the same neurone. The interaction of these two drugs was also studied. NA and IMI produced similar effects and a synergism between these two drugs was noted. These effects may be due to a blockade of the neurone uptake of NA by IMI. THEREis
considerable evidence to show that imipramine (IMT) antagonizes the uptake of noradrenaline (NA) by the postganglionic sympathetic innervation of various organs (GLOWINSKIand BALDESSARINI, 1966; IVERSEN, 1968). According to the “uptake hypothesis”, the main physiological function of NA uptake is to terminate the action of NA released from presynaptic adrenergic nerve terminals. Results that support this hypothesis have been obtained mainly in the peripheral nervous system (LANGERet al., 1967; IVERSEN,1968). Evidence for NA uptake in the CNS has been obtained by biochemical, histochemical and autoradiographic studies; in this case too, IMI seems to act as a blocker of NA uptake (GLOWINSKIand AXELROD, 1964; GLOWINSKI and BALDESSARINI, 1966; Ross and RENYI, 1967; CARLSS~Net al., 1968, 1969).
The present study concerns the effects of IMI on the impulse frequency of neurones in the CNS when this substance is released by microiontophoresis very close to the neurone under study. The effects of IMI were compared to those of NA applied microiontophoretically to the same neurone. The interaction of the two compounds was also studied. Results were obtained from 10 guinea pigs of either sex. Craniectomy was performed and the cerebellum was sucked out under ether anaesthesia. Wound margins and the pressure points of the head clamp were infiltrated with novocaine and the animal was immobilized with an i.p. injection of sincurarina (Farmitalia, SA Milano). The ether anaesthesia was then discontinued, and the animal was ventilated by a respiratory pump. For microiontophoresis, multibarrelled micropipettes (prepared according to BRADLEY et al., 1966) were used. One barrel of the micropipettes contained IMI hydrochloride, 7-IO%, pH 5, (Geigy, SA Milano); another contained NA bitartrate, IO%, pH 7, (KochLight Laboratories Ltd., Colnbrook, Buckinghamshire); a third barrel contained 1 M NaCl for monitoring current effects, and unitary discharges were recorded through a barrel containing 4 M NaCl. The micropipettes were inserted through the floor of the fourth ventricle. 661
662
G. L. AVANZINO,ROSAERMXRIO and C. ZUMMO
Fifty-one neurones, belonging mainly to the brain stem reticular formation, were studied Microiontophoretic appiication of NA by positive currents between 25 and 80 nA had inhibitory effects on 38 neurones, excitatory effects on 8, and no clear action on 5 neurones. Application of IMI with positive currents between 25 and 70 nA usually produced effects qualitatively similar to those of NA. IMI inhibited 26 of the neurones inhibited by NA and excited 5 of the neurones excited by NA. The effects of IMI were often weaker than those of NA (Figs. lA, B). Simultaneous application of NA and IMI produced longer or greater
A NA
-!!!L
N&I
30 -
f 20-
B
0
x)0
xx)
ma
-t FIG. 1. Effect of microiontophoretic application of noradrenaline (NA) and imipramine (IMI) on the luring frequency of 2 untts in the brain stem of a guinea pig. NA, applied at 40 nA for 30 set, had inhibitory (A) and excitatory (B) effects. IMI, applied at 30 nA for 40 see, weakly mimicked these actions of NA. The application of both substances together caused longer and more marked effects. In (A), current effects were tested by applying an amount of current equal to the total current used in applying NA and IMI. Axes: f, the total number of impulses every 5 set; t, the time in sec.
Imipramineand brain stem neurones
663
inhibition than was produced by the application of either compounds alone in 15 out of 18 neurones tested. Prolonged or increased excitation was seen in 2 neurones out of 6. When applied at 40-50 nA for 30 set, between 20 and 180 see before NA, IMI usually did not increase the NA effect (Fig. 2). When applied at 60-l 00 nA for 80-120 set i~~diately before NA, IMI occasionally depressed the response to NA,
NA
NA
30
f
20
10
~
FIG. 2. Effects of the application of nomdrenaline(NA}at 50 nA for 25 see and i~pr~ne (IMI}at 40nA for 3Osec.A second application of NA i2Osecafter the end of the IMI application had a similare&et to the first applicationof NA.
The current e%xts of simultaneous applications of NA and IMI were checked by passing the appropriate total current through the barrel containing 1 M NaCl (Fig. 1A) or by simultaneous application of current and NA, and of current and IMI. These results suggest that NA and IMI have qualitatively similar effects when applied by microiontophoresis to singIe neurones, and that there is synergism between these effects. It is suggested that these effects could be due to a blockade of the neuronal uptake of NA by IMI. The results so far obt~ned are not su~cient to determine whether the reduction of the effects of NA after prolonged applications of IMX are due to de~nsiti~tion of the NA effect or to an antagonistic action by IMI. REFERENCES BRADLEY,P, B., DHAWAN,B. N. and WOLSTENCROFT, J. IS. (1966). Pharmacological properties of cholinoceptive neurones in the medulla and pons of the cat. J. Physiol., Land. 183: 658-674. CARLWON, A., FUXE,K. and UNOERSTEDT, U. (1968). The effect of Imipramine on central 5 hydroxytryptamine neurons. J. Pharm. Pharmac. 20: 150-l 51. CARLSSON, A,, CORRODI, A., FUXE,K. and HBKFELT,T. (1969). ES?& of some antidepressant drugs on the depletion of intraneuronal brain catecholamine stores caused by 4 p dimethyl-men-tym~ne. Eur. J, Pharntac. 5: 367-373. GLOWIN~KI,
J. and AXELR~D,J. (1964). Inhibition of uptake of tritiated-noradrenaline in the intact rat brain by imipramine and structurafly related compounds. Nature, Load. 204: 1318-1319. ~~WINSKI, 3. and BALDESSARINI, R. J. (1966). Metallic of ~orepinep~ine in the Central Nervous System. Phar~ac. Rev. 18: 1201-1238.
664 fvmm,
G. L. AYANUNO, ROSA ERM~KXOand C. Zu~hno
L. It. (I%tQ. R&z #~~~~~~~~~~~ uptake in adrenergic n~~~srn~ss~~~. In: CTBA F&t. Symp. Adrenergic ~eur~transrni~~~n~ pp. 44-50. Churchill. London. LAwER, S. Z., DRASKOCZ~,R. R. and ~~~EL~NB~~~ U. f1967). Time course of the de~ekjpment of wpersensitivity to various amines in the nictitating membrane of the pithed cat after denervation or decentralization. 3. Phamac, exp. Tlrer, 157: 255-273. RCW, S, B, and RENYI, A. L. (1967). Inhibition ofthe uptake oftritiated catechotamines by antidepressant and related agents. Eur. J. Pharmac. 2: 181--186.