Effects of Neurokinin Receptor Antagonists on L-DOPA Induced Bladder Hyperactivity in Normal Conscious Rats

Vol. 154, 1548-1551, October 1995 Printed in U . S . A

EFFECTS OF NEUROKININ RECEPTOR ANTAGONISTS ON L-DOPA INDUCED BLADDER HYPERACTIVITY IN NORMAL CONSCIOUS RATS OSAMU ISHIZUKA, ANDERS MA'ITIASSON

AND

KARL-ERIK ANDERSSON*

From the Departments of UFology and Clinical Pharmacology, Lund University Hospital, Lund,Sweden ABSTRACT

Purpose: The urodynamic effects of intrathecal neurokinin (NK)receptor blockade on L-dopainduced bladder hyperactivity were investigated in a rat model. Materials and Methods: Continuous cystometry was performed in normal, conscious, female Sprague-Dawley rats. Results: In rats pretreated with intraperitoneal carbidopa 50 mg./kg., intraperitoneal L-dopa 50 mg./kg. caused bladder hyperactivity that could be attenuated by intrathecal administration of the NK, receptor selective antagonist SR 140,333(2 nmol.), whereas the NK, receptor selective antagonist SR 48,968(2 nmol.) failed to do so. Combination of SR 140,333(2 nmol.) and SR 48,968 (2 nmol.), which by itself decreased micturition pressure, practically abolished the L-dopainduced hyperactivity. Conclusions: The present results suggest that tachykinins, via stimulation of NK1 (and/or N&) receptors, are involved in L-dopa-induced bladder hyperactivity, most probably at the spinal level. "his implies tachykinin involvement in the supraspinal pathways that control the sacral parasympathetic center innervating the urinary bladder. It also implies that spinal NK receptors are a possible target for drugs aimed for elimination of bladder hyperactivity mediated via these pathways. KEYWORDS: bladder, rats, taehykinins, levodopa The sacral parasympathetic center innervating the urinary neurons localized inside the central nervous system." It can bladder is controlled via supraspinal pathways originating in therefore be conceived that tachykinins may also play a role the dorsolateral pontine tegmentum, including the locus co- in the supraspinal pathways originating in the dorsal poneruleus, which consists chiefly of norepinephrine-containing tine tegmentum, including the locus coeruleus, which control cells.1.z SillBn3.* showed in rats that the amine precursor the sacral micturition center. L-dopa given intrapentoneally, after peripheral inhibition of To clarify the roles of tachykinins in bladder hyperactivity the degradating enzyme dopa decarboxylase (by intraperitoneal presumed to originate in the dorsal pontine tegmentum, we carbidopa), produced a hyperactive urinary bladder response. studied in normal, conscious rats 1)the effects on the cysThis bladder action seemed to be elicitedlargely via stimulation tometrogram of intrapentoned L-dopa after pretreatment of dopamine receptors, most probably in the locu~coeruleus. with intraperitoneal carbidopa, 2 ) the changes of the effects Several peptides have been found in the central and of L-dopa produced by intrathecal administration of SR peripheral nervous systems innervating the lower urinary 140,333, a selective antagonist of NK, receptors12 and/or of tract. Among them, tachykinins (substance P and neuro- SR 48,968, a selective antagonist of NK, receptors.13.14 kinin A) and calcitonin gene-related peptide (CGRP) have been implied as mediators and/or modulators of capsaicinMATERIALS AND METHODS sensitive primary afferenta.6 It is known that both tachykinina and CGW are released from central endings of capsaAnimals. Female Sprague-Dawley rats, weighing 200 to icin-semitive primary afferent neurons in the rat and cat 215 g.9 were in The Pmbl spinal cor&,B*7and previous h c t i o n a l studies support the approved by the Animal Ethics Committee,University of Lund. Procedures: Bladder catheter implantation. Rats were view that the capsaicin-sensitive bladder afferenb are invalved in regulating the micturition reflex in the rat.s.s shar- anesthetized with ketamine (75 mg./kg. intramuscularly) technique and xylazine (15 mg./kg. intramuscularly). The abdomen was key et d.10demonstrated with double that 10 to 16% ofrat dorsal root ganglionic neurons, opened though a midline incision, and a polyethylene cathafferenta 6.om the bladder, contain substance p (sp). They eter (ClaY-Adms PE-50, Parsippany, New Jersey) implanted alsoshowed that sp-positiveneurons were notobserved in into the bladder through the dome as described previo~sly.'~ the dorsal ganglia of mpsaicin-pretreated It htrdhecal catheter implantation. An intrathecal catheter that capsaicin-sensitive fibers arising was implanted at the same time as the bladder catheter. A has been from the rat bladder may urn SP a8 an excitatory neurotrans- Polyethylene catheter (Clay-Adams PE-10) was inserted into mi&r to activate the second ne-n in the micturition reflex the subarachnoid space at the level of the L,S, spinal cord of the nerve fibers containing segments for intrathecal administration of drugs as depathway.8 Even if a large sp-mea u n o r e a c t i i t y (SP-LI)that can bobserved in the scribed in detail previous1y.l6 The injection sites in the spinal dorsal half of the spinal cord are of primary afferent origin, cord and the extent of dye distribution were confirmed by of the spinal hchykinin content seems to originate from injection of dye (methylene blue) in every animal at the end of the experiment. Accepted for publication A ril21, 1995. Intraperitoneal catheter implantation. An intraperito*Re uests for reprints: Jepadment of Clinical Pharmamkzy. neal catheter was implanted at the same time as the Lund Qniveraity Hospital, S-221 85 Lund, Sweden. ms wa8 by the Swedish Medical &search bladder catheter. A polyethylene catheter (Clay-Adam8 Council (no.6837 and 10399) and by the Medical Faculty of Lurid, PE-50) was inserted into the abdominal cavity for intraSweden. peritoneal administration of drugs. 1548

TACHYKININS AND SUPRASPINALLY INDUCED BLADDER HYPERACTMTY

1649

Cystometrical investigations. Cystometric investigations obtained before and after drug administration, Student's paired were performed without any anesthesia 3 days after bladder t test was used. One way factorial ANOVA was used for comcatheter implantation. The bladder catheter was connected parisons between L-dopa and L-dopa in the presence of SR a T-tube to a pressure transducer (P23 DC, Statham 140,333andor SR 48,968and was followed by Scheffe's F-test. Instrument Inc., Oxnard, California) and a microinjection A probability level of <5% was accepted as significant. pump (CMA 100, Carnegie Medicine AB, Solna, Sweden). m e conscious rat was placed, without any restraint, in a RESULTS metabolic cage which also permitted measurements of micEffects of intraperitoneal L-dopa. L-dopa, 50 mgJkg. (n = turition volumes by means of a fluid collector connected to a 61,decreased bladder capacity (p <0.001) and micturition Grass force displacement transducer (FT03 C, Grass Instru- volume (p <0.001) and increased basal pressure (p <0.01; ment Co., Quincy, Massachusetts). Room-temperaturesaline table 1;fig.). There were no changes in micturition pressure was infused into the bladder at a rate of 10 ml. per hour. and residual volume. The effeds of Gdopa appeared within Intravesical pressure and micturition volumes were recorded 10 minutes after the administration and continued for about continuously on a Grass polygraph (Model 7E, Grass Instru- 120 minutes. If the dose of Gdopa was increased to 100 ment Co.; recording speed 10 111111. per minute). Ten repro- mg./kg. (n = 12),a pronounced hyperactivity was produced ducible micturition cycles were recorded before drug admin- (data not shown). M o p a caused restless movements of the istration and used as baseline values. The following animals; this effect was dose-dependent. cystometric parameters were investigated17 micturition Effects of intmthecal SR 140,333 andlor SR 48,968.SR pressure (the maximum bladder pressure during micturi- 140,333(n = 6)or SR 48,968(n = 5), each in a dose of 2 nmol. 1 tion), bladder capacity (residual volume at the latest previous per rat, caused no changes in the cystometric parameters micturition plus volume of infused saline at the micturition), investigated (table 2). However, a combination of SR 140,333, micturition volume (volume of expelled urine) and residual 2 nmol. per rat and SR 48,968,2nmol. per rat (n = 5; table volume (bladder capacity minus micturition volume). The 10 2) decreased micturition pressure (p <0.05). There was no micturition cycles after the administration of Gdopa, in the change in other cystometric parameters and no abnormal presence or absence of SR 140,333 andor SR 48,968,were behavior. recorded. To study the effects of SR 140,333or SR 48,968,the Effects of intraperitod Mopa in the presence of intmtheml 2 micturition cycles showing the most pronounced changes SR 140,333 and forSR 48,968.In the presence of SR 140,333(n (increase or decrease) within 120 minutes after administra- = 6),2 nmol. per rat, the effects of L-dopa on bladder capacity tion of the drugs - were analyzed and compared with the (p C0.05)and micturition volume (p (0.05) were attenuated. baseline values. Also SR 48,968 (n = 5). 2 nmol. per rat, reduced the GdopaAdministration of drugs. SR 140,333,(S)1-12-[3-(3,4dichlo-induced effects on bladder capaciq (p cO.01). micturition volmphenyl~-l-~3-isopropoxyphenylacetyl)pipendin-3-yllethyl~ume (p <0.05)and basal pressure (p
B.C.

Gdopa 50 m g h g . before 62.5 ? 8.0 0.96 f 0.11 (n = 6) after 58.0 ? 3.2 0.36 -C 0.04*** Gdopa in the presence of intrathecal SR 140,333(2nmol.) before 96.9 f 16.8 0.942 0.11 50 mg&. (n = 6) after 91.62 17.1 0.69 f 0.05't Mops in the presence of intrathecal SR 48,968 (2nmol.) 50 mgJkg. before 95.6 2 7.1 0.91 2 0.15 (n = 6) after 83.1 f. 12.0 0.39 2 0.05** Mopa in the presence of intratheeal SR 140,333 (2nmol.) and SR 48,968 (2nm01.) before 77.0 2 9.3 0.96 2 0.17 60 m g k . (n = 6) after 68.82 7.7 0.74 2 0.lOt

M.V.

R.V.

B.P.

0.86 f.0.06 0.32 1+_ 0.03***

0.11 f 0.05 0.05 f 0.02

9.3 f 1.1 17.2 2 1.4"

0.89 f.0.12 0.61 f 0.06*t

0.06 2 0.02 0.09 2 0.02

8.1 2 1.1 11.4 2 1.W

0.84 f.0.15 0.34 f. 0.04'

0.08 2 0.02

10.4 f 3.9 15.3 f 4.6'

0.89 't 0.15 0.69 't 0.08W

0.07 2 0.03 0.052 0.02

0.05 2 0.02

9.8 2 2.5 9.6 2 2.Ott

M.P.: hcturitjon presaure (em.H,O);B.C.: bladder capacity ( d . h M.V.: micturition volume tml.); RV.:residual volume (d.h B.P.: basal preseurs (on.-0). Results are expressed as mean -C ~tandarderror of the mean. Comparisonsare made between before and after drug administratim:* p
WovA followed by Scheffe's F-test).

1550

TACHYKININS AND SUPRASPINALLY INDUCED BLADDER HYPERACTIVITY

I

-5

1

I

0

5

I

I

I

15

10

20

I

I

25

30

SR 140,333

h

1

I

-5

h

I

I

I

I

1

I

0

5

10

15

20

25

Time ( min ) Effects of Ldopa, administered intraperitoneally in dose of 50 mg./kg., on bladder pressure (BP; cm. H,O) and micturition volume (Mv, resence of intrathecally administered selective NK receptor antagonists. Carbidopa, 50 mg./kg., was given 15 minutes of Gdopa; B , effects of Gdopa in presence of SR 140,333,2nmol. per rat, and SR 48,968,2 nmol. per rat. Asterisk before L-dopa. A, e$& (*I denotes adjustment to baseline position. ml.) in absence and

TABLE2. Effects of intmthecal SR 140,333 andlor SR 48,968 on cystometric parameters in normal, conscious rats M.P.

B.C.

M.V.

R.V.

B.P.

SR 140,333 before 85.1 t 11.4 2 nmol. 0.96 t 0.10 1.11 2 0.10 0.14 t 0.02 8.0 2 1.2 1.05 t 0.09 0.96 t 0.08 (n = 6) after 80.8 2 7.9 0.092 0.02 7.52 0.9 SR 48,968 2 nmol. before 66.1 t 9.7 1.11 t 0.16 0.97 2 0.18 0.13 t 0.02 11.0 t 2.3 (n = 5) aRer 61.9 t 7.7 1.14 2 0.18 1.03 2 0.20 0.11 t 0.04 10.8 2 1.7 SR 140,333 and SR 48,968 2 nmol. before 84.2 2 13.3 1.11 2 0.18 0.90 t 0.11 0.21 2 0.09 12.1 ? 4.7 12.22 4.8 (n = 5) after 63.0 t 9.1' 1.19 t 0.26 0.97 2 0.14 0.23 t 0.13 M.P.:micturition pressure (em.HzO);B.C.: bladder capacity (ml.); M.V.: micturition volume (ml.); R.V.:residual volume (ml.); B.P.: basal pressure (cm. HzO). Results are expressed as mean t standard error of the mean. Comparison is made between before and aRer drug administration:* p ~0.05 (paired Student's two tailed t test).

the lateral marginal area of the dorsal horns and in the area of the sacral autonomic nucleus, which provide most of the efferent input to the urinary bladder.19 In previous investigations, performed in normal, urethaneanesthetized animals, different types of micturition-related reflexes have been elicited: 1) a supraspinal chemonociceptive vesicovesical reflex produced by topical application of capsaicin onto the dome of the urinary bladder, 2) a supraspinal mechanoreceptive reflex evoked by saline filling of the urinary bladder, and 3) a spinal somatovesical mechanonociceptive reflex evoked by perineal pinching.20 These studies, in which several NK, receptor selective antagonists were used, provided evidence for the involvement of tachykinins, through spinal NK, receptors, in the activation of reflexes produced by topical application of capsaicin to the serosal side of the bladder. A modulatory influence on bladder capacity elicited by distension was also suggested. Magnan et

al.,*l in studying the effects of intrathecally administered

NK, and NK, antagonists on xylene-induced hyperactivity in urethane-anesthetized animals, arrived at similar conclusions, suggesting that NK,receptors at the spinal level are involved in the bladder response to chemonociceptive stimuli. However, they found no evidence in support of involvement of spinal NKl (or NKJ receptors in regulation of normal micturition. Ishizuka et a1.22showed that the NK, receptor selective antagonist, R P 67,580, when given intrathecally to normal animals, reduced micturition pressure, and in 1of 7 animals produced dribbling incontinence due to urinary retention. In this animal, the effect was reversible. Since dribbling incontinence was not produced by intrathecal saline or by RP 68,651, the inactive enantiomer of RP 67,580, the finding wa8 interpreted as an effect of the NK, antagonist. In animals with bladder outflow obstruction, a condition known to facilitate bladder reflex mechanisms,23 RP 67,580 induced

TACHYKININS AND SUPRASPINALLY INDUCED BLADDER HYPERACTIVITY

nary retention in 5 of 7 animals.22 The failure of the highly potent nonpeptide NK, receptor antagonist SR 48,968 to affect micturition when given intrathecally to both normal rats and animals with outflow obstruction seemed to exclude spinal involvement of NK, receptors in the micturition reflexes induced by filling. "his interpretation is in line with the fact that only a few studies have detected NK,receptors in the spinal cord.24 In the present study, the selective NK, antagonist SR 140,333tended to decrease the micturition pressure, but this effect was not significant. The selective NK,antagonist SR 48,968also was without a significant effect. The only effect of the combination of SR 140,333and SR 48,968was a decrease in micturition pressure. This may be taken as further support of the view that tachykinins, if involved in the micturition reflex induced by bladder filling in normal animals, are of minor importance. After pretreatment with intraperitoneal carbidopa, intraperitoneal L-dopa will stimulate micturition or induce bladder hyperactivity in anesthetized rats.3.25 "his was also the case in normal, conscious rats. Sillen et al.3 suggested that the L-dopa induced hyperactivity is elicited chiefly via stimulation of central dopaminergic receptors, because the effects were abolished by prior intraperitoneal administration of the centrally acting dopamine receptor blocker, spiroperidol. G dopa probably acts on the dopaminergic receptors in the locus coeruleus. From the locus coeruleus, which has been implicated in the supraspinal control of micturition,26 noradrenergic neurons supply sympathetic and parasympathetic nuclei in the lumbosacral spinal cord. These bulbospind noradrenergic pathways may be of importance for micturition. Elam et a1.26 showed that distension of the rat urinary bladder and concomitant increase in bladder pressure was associated with a n increase in the firing rate of noradrenaline neurons in the locus coeruleus. In the anesthetized cat, electrical stimulation of the locus coeruleus induced bladder contractions that were blocked by intrathecal administration of prazosin,27 suggesting involvement of a-adrenoceptors. However, Sillen et al.4 found evidence also for the existence of a nonadrenergic, noncholinergic pathway between the pontine and spinal micturition centers. It has been suggested that the peripheral mediation of the bladder hyperactivity initiated by L-dopa is propagated via the pelvic nerves.= The present results suggest that tachykinins, via stimulation of NK, (and/or NK,) receptors, are involved in L-dopa induced bladder hyperactivity, most probably at the spinal level. This means that tachykinins may play a role for the supraspinal pathways originating in the dorsal pontine tegmentum, including the locus coeruleus, that control the sacral parasympathetic center innervating the urinary bladder. It also implies that spinal NK receptors are a possible target for drugs aimed at eliminating of bladder hyperactivity mediated via these pathways. Acknowledgement. The generous giR of tachykinin antagonists by Dr. A. Lecci is gratefully acknowledged. REFERENCES

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