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Role of central tachykinin peptides in cardiovascular regulation in rats
Brain Research, 528 (1990) 231-237 Elsevier
231
BRES 15873
Role of central tachykinin peptides in cardiovascular regulation in rats Yukio Takano 1, Akira Nagashima 1, Tetsuya Hagio 1, Kayoko Tateishi 2 and Hiro-o Kamiya 1 t Department of Pharmacology, Faculty of Pharmaceutical Sciences and 2The First Department of Biochemistry, School of Medicine, Fukuoka University, Fukuoka (Japan) (Accepted 20 March 1990) Key words: Substance P; Tachykinin peptide; Neurokinin B; Cardiovascular regulation; Vasopressin; Blood pressure
The mechanisms of action of tachykinin peptides thought to be involved in central cardiovascular regulation were examined. Intracerebroventricular injections (i.c.v.) of tachykinin peptides caused dose-dependent increases in blood pressure and heart rate. The pressor responses to substance P (SP) (10/~g, i.c.v.) and neurokinin A (NKA) (10/tg, i.c.v.) were blocked by peripheral administration of pentolinium or phentolamine, and partially attenuated by adrenalectomy. In contrast, the only initial pressor response to the neurokinin B (NKB) analogue senktide (10 pg, i.c.v.) was blocked by pentolinium or phentolamine. The pressor response to senktide was inhibited by pretreatment with a vasopressin V 1 receptor antagonist (d(CH2)5OMe(Tyr)AVP) (10/~g/kg, i.v.), and senktide (10 jug, i.c.v.) caused an increase in plasma vasopressin level. However, the vasopressin antagonist did not influence the SP- and NKA-induced pressor responses. These results suggest that central SP and NKA increase the blood pressure and heart rate via sympathetic nerve activity, whereas central NKB increases the blood pressure mainly via release of vasopressin from the hypothalamus. INTRODUC~JON Central neural circuits play important roles in cardiovascular control by regulating autonomic functions or vasopressin release from the hypothalamus. Recently, m a n y neuropeptides that seem to be involved in central blood pressure control have been discovered. These neuropeptides have become of interest as endogenous substances regulating blood pressure like neurotransmitters, such as catecholamine, acetylcholine, serotonin, histamine and y-aminobutyric acid ( G A B A ) . Although much information has been obtained on these peptides (for reviews, see refs. 8, 27, 29), their effects on the cardiovascular system are still not clearly understood. Substance P (SP) is a m e m b e r of the tachykinin family of bioactive peptides, which includes neurokinin A ( N K A ) , neurokinin B (NKB), neuropeptide K (NPK) and neuropeptide y (NPy) (for reviews, see refs. 6, 18, 22). SP, N K A , N P K and NPy are derived from the preprotachykinin A gene, whereas NKB is derived from the preprotachykinin B gene 15-17'25'26. From cardiovascular findings, SP has become of interest as a neuropeptide regulating the blood pressure in the ventral medulla 5" 13,14,20,33 nucleus tractus solitarii (NTS) 4'10-12'21'23'36-38 and spinal cord 9'34'35. Intracerebroventricular injection (i.c.v.) of SP increases the blood pressure. This pressor response to SP seems to be regulated by increase in sympathetic nerve activity, without participation of vaso-
pressin 7"39'41'42'44. Previously, we found that the NKB analogue senktide (i.c.v.) also causes increase in blood pressure, and that its action seemed to be mediated by vasopressin secreted from the hypothalamus, because blockade of peripheral vascular vasopressin receptors inhibited the pressor response 24. However, little is known about the mechanisms of central cardiovascular regulation induced by tachykinin peptides. In the present study, we examined the central mechanisms of action of tachykinin peptides thought to be involved in cardiovascular regulation. First, we studied the effects of sympathetic blockade and a vasopressin antagonist on tachykinin-induced central pressor responses. Then, we examined the effects of tachykinin peptides (i.c.v.) on plasma vasopressin levels. MATERIALS AND METHODS Animals Male Wistar rats (250-300 g) were purchased from Kyudo (Kumamoto, Japan). The animals were kept in a room at 20-25 °C with a 12 h light-dark cycle (light on at 07.00 h) and were given free access to commercial food and tap water. Cardiovascular response Experiments on cardiovascular responses were performed as described previously 23,24. Rats were anesthetized with urethane (1 g/kg i.p.). Then one catheter was inserted into the right femoral artery for recording the blood pressure and heart rate, and another into the left femoral vein for administration of drugs. The body temperature was maintained at 37-38 °C with a heating pad. The
Correspondence: Y. Takano, Department of Pharmacology, Faculty of Pharmaceutical Sciences, Fukuoka University, Fukuoka 814-01, Japan. 0006-8993/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)
232 systolic blood pressure was maintained above 99 mm Hg by infusion of 4% Ficoll 70 (Nacalai Chemicals, Japan) in saline solution. Each rat was placed in a stereotaxic apparatus. A guide cannula (21-gauge stainless steel) was inserted into the lateral brain ventricle. The adrenal glands were removed bilaterally from some animals. A stabilization period of 30 min was allowed before administration of drugs. Peptides were dissolved in artificial cerebrospinal fluid (ACSF; in raM): NaCI 128, KCI 3, CaCI2 1.2, MgCI2 0.8, NaH2PO 4 0.25, NaHCO3 21 and glucose 3.4; pH 7.4) and 10pl volumes of solutions were injected manually over a period of 1 min with a Hamilton microsyringe. Injection of ACSF (i.c.v.) did not cause any detectable change in blood pressure or heart rate. After each experiment, Malachite-green was injected to confirm the site of injection.
Plasma vasopressin level Rats were decapitated 5 or 20 min after i.c.v, injections, and
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Fig. 1 summarizes the effects of intracerebroventricular injections (i.c.v.) of various tachykinin peptides on the mean arterial blood pressure and heart rate. The central pressor responses to the tachykinin peptides were dose-dependent, reaching maxima 4-6 rain after injections of the peptides and then persisting for at least 40 rain.
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SP and NKA were obtained from the Peptide Institute (Osaka, Japan). [1-(fl-fl-Cyclopentamethylene propionic acid, 2-(O-methyl)tyrosine] ArgS-vasopressin (d(CH2)5OMe(Tyr)AVP), [Pro7]-NKB, (suc-[Asp6,Me-PheS]-substance P(6-11) (senktide), [(AspS'6,Me PheS]substance P(5-11) (NHz-senktide) were from Peninsula (California, U.S.A). NPy was a gift from Dr. J.E. Krause (Washington University, St. Louis, U.S.A.). Data are expressed as means + S.E.M.s. Statistical significance was determined by the two-tailed Student's t-test, or the Dunnett's multiple comparison test.
Cardiovascular effects of tachykinin peptides 6. c. v.)
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Peptides and statistical analysis
RESULTS
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blood samples were collected in chilled tubes containing 1.2 mg EDTA-2Na. The plasma was stored at -70 °C, until just before use. Plasma samples (200/A) were mixed with an equal volume of 0.1 N HCI and passed through a Sep-Pac Cls cartridge (Millipore Waters, MA). The Sep-Pac C18 cartridge was washed with 4% acetic acid and then adsorbed vasopressin was eluted with 1.5 ml of methanol, and dried under nitrogen gas. Radioimmunoassay (RIA) was performed with a vasopressin-RIA kit (Mitsubishi-Yuka, Tokyo, Japan). The antiserum showed less than 0.01% cross-reactivity with oxytocin. The sensitivity of the assay was 0.063 pg/tube.
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pressure a n d h e a r t rate induced by tacbykinin peptides (i.c.v.) at different doses, (A) substance P, (B) neurokinin A, and (C) senktide. Values are means _+ S.E.M.s for 6-10 rats. Fig. 1. T i m e course o f the responses in b l o o d
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To determine the involvement of tachykinin peptides in the sympathetic nervous system, we tested the effects of sympathetic blocking agents on the cardiovascular responses. The ganglionic blocker, pentolinium, was administered i.v. 10 min before injections of tachykinin peptides (i.c.v.). Pentolinium (10 mg/kg, i.v.) reduced the basal blood pressure. Pretreatment with pentolinium significantly inhibited the increases in blood pressure and heart rate elicited by SP and NKA. Although pentolinium blocked the earlier pressor response induced by the NKB analogue senktide, the later blood pressure after injection of senktide was increased under sympathetic ganglionic blockade (Fig. 2). The effects of peripheral a- and fl-adrenoceptor blockades are summarized in Table I. Administration of an a-adrenoceptor antagonist, phentolamine (10 mg/kg, i.v.), reduced the basal blood pressure, and slightly reduced the basal heart rate. Under peripheral aadrenoceptor blockade, the central pressor responses induced by SP and NKA were blocked. In contrast, pretreatment with phentolamine blocked only the initial pressor response induced by senktide (i.c.v.) (Table I). Administration of a fl-adrenoceptor antagonist, propranolol (1 mg/kg, i.v.), decreased in heart rate. Under peripheral fl-adrenoceptor blockade, the pressor response and tachycardiac action induced by SP (10 pg, i.c.v.) were inhibited. Pretreatment with propranolol inhibited tachycardia induced by NKA or senktide, but did not change the NKA-induced pressor response. Propranolol had no effect on the hypertensive effect of senktide (10/~g, i.c.v.) (Table I).
Effect of adrenalectomy on cardiovascular responses to tachykinin peptides (i.c. v. ) Bilateral adrenalectomy had no effect on the basal blood pressure or heart rate. Injection of SP or NKA increased the blood pressure and heart rate in both the sham-operated control group and the adrenalectomized group, but sustained pressor responses (20-40 min after i.c.v, injections) were significantly less in the adrenolectomized group than in the control group (Table II). Adrenalectomy did not block the senktide-induced pressor response and had no effect on the tachycardia induced by tachykinin peptides (i.c.v.) (Table II).
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234 TABLE I Effects of a- and fl-adrenoreceptor blocking agents, and vasopressin V1 antagonist (i.v.) on the cardiovascular responses to tachykinin peptides (i.c.v.) Data are mean _+S.E.M. n
Substance P (10/~g, i.c.v.) Saline 11 Phentolamine (10 mg/kg, i.v.) 8 Propranolol (1 mg/kg, i.v.) 8 Vl-antagonist (10 mg/kg, i.v.) 7 Neurokinin A (10/~g, i.c.v.) Saline 13 Phentolamine (10 mg/kg, i.v.) 6 Propranoloi (1 mg/kg, i,v.) 6 Vl-antagonist (10/~g/kg, i.v.) 6 Senktide (10/tg, i.c.v.) Saline 6 Phentolamine (10mg/kg, i.v.) 7 Propranoloi (1 mg/kg, i.v.) 10 Vl-antagonist (10~g/kg, i.v. ) 6
Basal MAP
A M A P (mm Hg) a
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5 rain
20 min
88.4_+6.70 46.6 _+1.24** 97.1_+3.26 76.9_+5.99
+23.2+2.09 +4.25 + 1.24"* +12.6 _+2.26** +19.6_+1.51
+12.8-+2.08 +1.00 + 1.45"* +0.50 _+2.75** +13.0_+2.48
89.8_+5.09 49.7_+3.16"* 97.3_+4.74 77.1_+4.58
+18.5_+1.73 +7.67 _+0.76* +23.7 _+3.81 +17.3 _+4 . 5 0
90.2+ 8.81 46.6_+2.89** 98.7_+4.70 81.1_+3.11
+15.3_+2.14 +6.00_+1.36" +14.6_+2.52 +5.17 _+1.76"
AHR (beats~rain)a 5 min
20 rain
422.4+13.1 422.3 _+19.1 321.4 _+6.36** 423.7_+15.2
+17.3-+5.15 +26.6 _+8.52 +19.8 + 5.07 +31.7_+12.7
+34.0 +25.9 +1.63 +17.1
+12.3_+1.80 +5.17 _+1.90 + 13.7 + 3.89 +14.3 _+2.21
441.0_+11.0 388.3 _+23.2* 295.2 + 12.3"* 426.6 _+7.33
+43.9+6.30 + 14,4 _+4.37 +38.2 _+ 15.4 +31.6 _+ 10.4 + 14.8 + 4.03* +14.8 _+4.03 +27.3 _+9.13 +20.8 + 8.29
+4.33+4.00 +7.71_+1.51 +11.1_+2.51 -18.8 _+2.36**
452.2_+21.6 418.0_+9.31 323.9+7.25** 410.1 _+15.3
+16.7_+6.29 +16.1_+3.78 +3.80+4.75 +19.2 _+6.31
_+5.50 _+6.47 _+5.25** + 6.22
+14.7 -+ 11.1 +5.57 -+ 6.21 +6.60 + 4.99 + 15.7 _+6.38
*P < 0.05 and **P < 0.01 vs corresponding value for saline control by the Dunnett's test. aChanges in mean arterial blood pressure (MAP) and heart rate (HR) at 5 and 20 min after i.c.v, administration of the peptides. Effects o f vasopressin antagonist on cardiovascular responses to tachykinin peptides (i.c.v.) To d e t e r m i n e whether vasopressin receptors in the peripheral vasculature are involved in the pressor responses observed after injections (i.c.v.) of tachykinin peptides, we injected the vasopressin V 1 receptor antagonist (i.v.) 10 min before tachykinin peptides. The vasopressin antagonist (10/~g/kg, i.v.) did not cause any detectable changes in either the blood pressure or the heart rate. P r e t r e a t m e n t with the vasopressin antagonist completely inhibited the sustained hypertension 4 - 4 0 min after injection induced by senktide (10 ~g, i.c.v.), and
reduced the blood pressure to below the basal level after 20 rain (Table I). It also inhibited the hypertension induced by other NKB analogues, such as [Pro7]-NKB and NH2-senktide (data not shown). It did not, however, significantly affect the increases in blood pressure induced by SP and N K A (Table I). Effects o f tachykinin peptides (i. c. v.) on p l a s m a vasopressin level Fig. 3 shows the plasma vasopressin levels 5 and 20 min after injections (i.c.v.) of tachykinin peptides. Injection of senktide (10/~g, i.c.v.) caused a m a r k e d increase in the
TABLE II Effects of bilateral adrenalectomy on the cardiovascular responses to tachykinin peptides (i.c. v. ) Data are mean + S.E.M. n
BasalMAP
A M A P (mm Hg) a 5 rain
20 min
BasalHR 40 min
Substance P (10~g, i.c.v.) Sham 7 85.8+4.90 Adrenalectomy 8 82.3+4.98
+17.9_+3.43 +9.14_+5.00 +15,4_+2.74 -0.38_+2.20
Neurokinin A (10/~g, i.c.v.) Sham 8 84.3+5.87 Adrenalectomy 8 86.6+8.04
+24.9-+3.00 +19.9-+3.93 +18.1+3.90 428.0+11.0 +28.0_+4.17 -6.25_+7.64** -9.88_+8.55** 420.8_+21.3
Senktide (10/~g, i.c.v.) Sham 8 88.2+2.85 Adrenalectomy 8 84.3_+4.68
+16.9-+2.26 +3.44-+3.21 +12.1_+3.51 -1.11_+5.62
AHR (beats~rain)~ 5 rain
20 min
40 rain
+6.14_+4.66 425.4_+13.3 +41.7_+7.29 +31.0_+12.2 +23.6_+10.6 -8.13_+4.32" 415.9_+13.7 +36.0_+7,21 +14.8_+4.46 +12.8_+5.88 +29.3+6.01 +45.8+11.8
+25.9_+9.22 +21.3_+12.2 +21.6_+13.9 +28.8_+8.73
+1.33+2.30 436.7-+10.1 +17.0_+4.86 +8.67_+8.33 +8.78+4.16 +0.78_+3.82 430.7_+12.0 +31.2+8.57 -2,11_+8.32 +5.00_+13.0
*P < 0.05 and **P < 0.01 vs corresponding value for sham-operated control by the two-tailed Student's t-test. aChanges in mean arterial blood pressure (MAP) and heart rate (HR) at 5, 20 and 40 min after i.c.v, administration of the peptides.
235 Angiotensin II 10ug (ICV)
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Angiotensin II in the brain stimulates the releases of vasopressin, oxytocin and adrenocorticotropic hormone (ACTH), and induces increase in blood pressure. To determine the contribution of the central angiotensin II system to the hypertension induced by senktide, we examined the effect of pretreatment with the angiotensin II receptor antagonist saralasin on the senktide-induced pressor response. Injection of saralasin (10 big, i.c.v.) increased the basal blood pressure, which returned to the basal level 10 min after the injection. Pretreatment with saralasin (i.c.v.) completely abolished the pressor response induced by angiotensin II (10/~g, i.c.v.) but did not influence that induced by senktide (Fig. 4).
2ok
DISCUSSION
There are several reports that the central pressor response to SP seems to be mediated by the sympathetic nervous system, without participation of vasopressin in conscious rats 41"42"44. However, there are few reports of studies on the central roles of other tachykinin peptides, probably partly because of the poor solubility of NKB in water and the absence of selective ligands for the tachykinin receptor subtype. Recently, selective NKB receptor ligands, senktide, NHE-senktide and [Pro7] NKB, have been developed 19'45. The present results
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the pressor responses after injections of (A) angiotensin II and (B) senktide. Values are means _+ S.E.M.s for 4 rats. *P < 0.05, **P < 0.01 and ***P < 0.005 vs corresponding value for ACSF control by the two-tailed Student's t-test.
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236 indicate a possible mechanism of central cardiovascular regulation by tachykinin peptides. Injection (i.c.v.) of the NKB-analogue senktide caused dose-dependent increase in blood pressure, and this response was inhibited by pretreatment (i.v.) with vasopressin V 1 receptor antagonist. In addition, senktide caused a marked increase in the plasma vasopressin level. Very recently Polidori et al. 28 reported similar increases of the plasma vasopressin level by NKB analogues. These findings suggest that the central cardiovascular effects of NKB analogues are mediated mainly by vasopressin. Presumably, treatment with an NKB analogue (i.c.v.) stimulates the release of vasopressin from the paraventricular nucleus (PVN) and/or supraoptic nucleus (SON) of the hypothalamus through the NKB receptor (NK-3 subtype) (Fig. 5), because we found previously the PVN and SON had higher contents of NKB-like immunoreactivity than other areas 24. Moreover high densities of [3H]NKB binding sites are present in these areas 2'3°. But the initial pressor responses to NKB analogues were inhibited by pretreatment (i.v.) with pentolinium or phentolamine, suggesting that the sympathetic nervous system initially contributes to the NKB analogue-induced pressor response. Injection of angiotensin II into the lateral brain ventricle (i.c.v.) is reported to cause an increase in blood pressure by stimulating release of vasopressin 31. However, this central angiotensin II system is not involved in the senktide-induced pressor response, because the angiotensin II receptor antagonist saralasin (i.c.v.) did not influence the senktide-induced pressor response (Fig. 4). It is unknown why the senktide-induced pressor response was reduced below the basal level after pretreatment with the vasopressin V 1 receptor antagonist (Table I). One possible explanation of this finding is that vasopressin released into the plasma by senktide (i.c.v.) stimulates not only the vascular vasopressin V 1 receptor but also the vasopressin receptor subtype in the area postrema (AP). Recently vasopressin was found to increase the sensitivity of the baroreceptor reflex through the vasopressin receptor subtype in the area postrema 1' 3,32,40,43 Therefore, under blockade of the vascular
vasopressin V 1 receptor, the vasopressin released by senktide may enhance the baroreceptor reflex inhibition of sympathetic nerve activity through the vasopressin receptor in the area postrema (Fig. 5). This idea is supported by the finding that the sustained blood pressure induced by senktide was enhanced under sympathetic ganglionic blockade (Fig. 2). The NKA-induced pressor response was reduced by blockade of the sympathetic nervous system, and NKA did not alter the plasma vasopressin level. These findings suggest that the pressor responses to NKA as well as SP are mediated by the sympathetic nervous system. Another tachykinin peptide derived from the preprotachykinin A gene, NP~, (i.c.v.), also increased the blood pressure without influencing the plasma vasopressin level (unpublished data). Since the increases in heart rate induced by SP and NKA are blocked by the peripheral administration of pentolinium and propranolol, the heart rate responses to SP and NKA also seem to be mediated by the sympathetic nervous system. Removal of the adrenal glands attenuated the sustained pressor responses induced by SP and NKA. These findings suggest that SP and NKA activate sympathetic nerves to the vasculature and adrenal catecholamine secretion. Thus the initial pressor responses to SP and NKA may be due to the sympathetically mediated vasoconstriction, and the sustained pressor responses to adrenal catecholamine secretion. Fig. 5 is a diagram summarizing our proposal on the central cardiovascular regulations by tachykinin peptides. We postulate that central SP and NKA, derived from the preprotachykinin A gene, increase the blood pressure and heart rate via stimulation of sympathetic nerve activity, whereas central NKB, derived from the preprotachykinin B gene, increases the blood pressure via release of vasopressin from the hypothalamus. Acknowledgements. This work was supported in part by a grant from the Suzuken Memorial Foundation. We are grateful to Dr. A.D. Loewy (Washington University, St. Louis, MO) for helpful discussion. We thank T. Akiyama, K. Hasegawa, and K. Takakura for technical assistance.
REFERENCES 1 Applegate, R.J., Hasser, E.M. and Bishop, V.S., Vagal cold block in area postrema-lesioned dogs: interaction of vasopressin and sympathetic nervous system, Am. J. Physiol., 252 (1987) H135-H141. 2 Bergstrom, L., Torrens, Y., Saffroy, M., Beaujouan, J.C., Lavielle, S., Chassaing, G., Morgat, J.L., Glowinski, J. and Marquet, A., [aH]Neurokinin B and 125I-BoltonHunter eledoisin label identical tachykinin binding sites in the rat brain, J. Neurochem., 48 (1987) 125-133. 3 Bishop, V.S, Hasser, E.M. and Undesser, K.E, Vasopressin and sympathetic nerve activity: involvement of the area postrema. In J.E Backley and C.M. Ferrario (Eds.), Brain Peptides and
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6 7
Catecholamines in Cardiovascular Regulation, Raven, New York, 1987, pp. 373-382. Carter, D.A. and Lightman, S.L., Substance P microinjections into the nucleus tractus solitarius elicit a pressor response in capsaicin-treated rats, Neurosci. Len., 43 (1983) 253-257. Charlton, C.G. and Helke, C.J., Substance P-containing medullary projections to the intermediolateral cell column: identification with retrogradely transported rhodamine-labeled latex microspheres and immunohistochemistry, Brain Research, 418 (1987) 245-254. Erspamer, V., The tachykinin peptide family, Trends Neurosci., 4 (1981) 267-269. Fuxe, K., Agnati, L.E, Rosell, S., Haffstrand, A., Foiker, K., Lundberg, J.M., Andersson, K. and H0kfelt, T., Vasopressin
237 effects of substance P and C-terminal sequence after intracisternal injection to a-chloralose anesthetized rats: blockade by a substance P antagonist, Fur. J. Pharmacol., 77 (1982) 171-176. 8 Gardiner, S.M. and Bennett, T., Brain neuropeptides: actions on central cardiovascular control mechanisms, Brain Res. Rev., 14 (1989) 79-116. 9 Gilbey, M.E, McKenna, K.E. and Schramm, L.P., Effects of substance P on sympathetic preganglionic neurones, Neurosci. Lett., 41 (1983) 157-159. 10 Gillis, R.A., Helke, C.J., Hamilton, B.L., Norman, W.P. and Jacobowitz, D.M., Evidence that substance P is a neurotransmitter of baro- and chemoreceptor afferents in the nucleus tractus solitarius, Brain Research, 181 (1980) 476-481. 11 Haeusler, G. and Osterwalder, R., Evidence suggesting a transmitter or neuromodulatory role for substance P at the first synapse of the baroreceptor reflex, Naunyn-Schmiedeberg's Arch. PharmacoL, 314 (1980) 111-121. 12 Helke, C.J., O'Donohue, T.L. and Jacobowitz, D.M., Substance P as a baro- and chemoreceptor afferent neurotransmitter: immunocytochemical and neurochemical evidence in the rat, Peptides, 1 (1980) 1-9. 13 Helke, C.J., Neii, J.J., Massari, V.J. and Loewy, A.D., Substance P neurons project from the ventral medulla to the intermediolateral cell column and ventral horn in the rat, Brain Research, 243 (1982) 147-152. 14 Helke, C.J., Shults, C.W., Chase, T.N. and O'Donohue, T.L., Autoradiographic localization of substance P receptors in rat medulla: effect of vagotomy and nodose ganglionectomy, Neuroscience, 12 (1984) 215-223. 15 Kawaguchi, Y., Hoshimaru, M., Nawa, H. and Nakanishi, S., Sequence analysis of cloned cDNA for rat substance P precursor: existence of a third substance P precursor, Biochem. Biophys. Res. Commun., 139 (1986) 1040-1046. 16 Kotani, H., Hoshimura, M., Nawa, H. and Nakanishi, S., Structure and gene organization of bovine neuromedin K precursor, Proc. Natl. Acad. Sci. U.S.A., 83 (1986) 7074-7078. 17 Krause, J.E., Chirigwin, J.M., Carter, M.S., Xu, Z.S. and Hershey, A.D., Three rat preprotachykinin mRNAs encode the neuropeptides substance P and neurokinin A, Proc. Natl. Acad. Sci. U.S.A., 84 (1987) 882-885. 18 Krause, J.E., MacDonald, M.R. and Takeda, Y., The polyprotein nature of substance P precursors, BioEssays, 10 (1989) 62-69. 19 Laufer, R., Gilon, C., Chorev, M. and Selinger, Z., Characterization of a neurokinin B receptor site in rat brain using a highly selective radio ligand, J. Biol. Chem., 261 (1986) 10257-10263. 20 Loewy, A.D. and Sawyer, W.B. Substance P antagonist inhibits vasomotor responses elicited from ventral medulla in rat, Brain Research, 245 (1982) 379-383. 21 Lukovic, L., De Jong, W. and De Wied, D., Cardiovascular effects of substance P and capsaicin microinjected into the nucleus tractus solitarii of rat, Brain Research, 422 (1987) 312-318. 22 Maggio, J.M., Tachykinins, Annu. Rev. Neurosci., 11 (1989) 13-28. 23 Nagashima, A., Takano, Y., Tateishi, K., Matsuoka, Y., Hamaoka, T. and Kamiya, H., Cardiovascular roles of tachykinin peptides in the nucleus tractus solitarii of rats, Brain Research, 487 (1989) 392-396. 24 Nagashima, A., Takano, Y., Tateishi, K., Matsuoka, Y., Hamaoka, T. and Kamiya, H., Central pressor actions of neurokinin B: increases in neurokinin B contents in discrete nuclei in spontaneously hypertensive rats, Brain Research, 499 (1989) 198--203. 25 Nakanishi, S., Substance P precursor and kininogen: their structures, gene originations, and regulation, Physiol. Rev., 67 (1987) 227-241. 26 Nawa, H., Hirose, T., Takashima, H., Inayama, S. and Nakanishi, S., Nucleotide sequences of cloned cDNAs for two types of bovine brain, substance P precursor, Nature, 306 (1983) 32-36. 27 Philippu, A., Regulation of blood pressure by central neuro-
transmitter and neuropeptides, Rev. Physiol. Biochem. Pharmacol., 111 (1988) 1-115. 28 Polidori, C., Saija, A., Perfumi, M., Costa, G., de Caro, G. and Massi, M., Vasopressin release induced by intracranial injection of tachykinins is due to activation of central neurokinin-3 receptor, Neurosci. Lett., 103 (1989) 320-325. 29 Reid, J.L. and Rubin, EC., Peptides and central regulation of the circulation, Physiol. Rev., 67 (1987) 725-749. 30 Saffroy, M., Beaujouan, J.C., Torrens, Y., Besseyre, J., Bergstrom, L. and Glowinski, J., Localization of tachykinin binding sites (NK1, NK2, NK3 ligands) in the rat brain, Peptides, 9 (1988) 227-241. 31 Sever, W.B. and Daniels-Severs, A.E., Effects of angiotensin on the central nervous system, Pharmacol. Rev., 25 (1973) 415-449. 32 Suzuki, S., Takeshiro, A., Imaizumi, T., Hirooka, Y., Yoshida, M., Ando, S. and Nakamura, M., Central nervous system mechanisms involved in inhibition of renal sympathetic nerve activity induced by arginine vasopressin, Circ. Res., 65 (1989) 1390-1399. 33 Takano, Y., Martin, J.E., Leeman, S.E. and Loewy, A.D., Substance P immunorcactivity released from rat spinal cord after kainic acid excitation of the ventral medulla oblongata: a correlation with increased blood pressure, Brain Research, 291 (1984) 168-172. 34 Takano, Y. and Loewy, A.D., Reduction of [3H]substance P binding in the intermediolateral cell column after sympathectomy, Brain Research, 333 (1985) 193-196. 35 Takano, Y., Sawyer, W.B. and Loewy, A.D., Substance P mechanisms of the spinal cord related to vasomotor tone in the spontaneously hypertensive rat, Brain Research, 334 (1985) 106-116. 36 Takano, Y., Nagashima, A., Masui, H., Kuromizu, K. and Kamiya, H., Distribution of substance K (neurokinin A) in the brain and peripheral tissues of rats, Brain Research, 369 (1986) 400-404. 37 Takano, Y., Nagashima, A., Kamiya, H., Kurosawa, M. and Sato, A., Well-maintained reflex responses of sympathetic nerve activity to stimulation of barorcceptor, chemoreceptor and cutaneous mechanoreceptors in neonatal capsaicin-treated rats, Brain Research, 455 (1988) 188-192. 38 Talman, W.T. and Reis, D.J., Baroreflex actions of substance P microinjected into the nucleus tractus solitarii in rat: a consequence of local distortion, Brain Research, 220 (1981) 402-407. 39 Trimarchi, G.R., Glisson, W.G., Thomson, W.M., Van Lingen, J. and Buccafusco, J.J., Cholinergic neurons and the cardiovascular response produced by central injection of substance P in the normotensive rat, Life Sci., 39 (1986) 1579-1588. 40 Undesser, K.P., Hasser, E.M., Haywood, J.R., Johnson, A.K. and Bishop, V.S., Interactions of vasopressin with area postrema in arterial baroreflex function in conscious rabbits, Circ. Res., 56 (1985) 410-417. 41 Unger, T., Rascher, W., Schuster, C., Pavlovitch, R., Schomig, A., Dietz, R. and Ganten, D., Central blood pressure effects of substance P and angiotensin II: role of the sympathetic nervous system and vasopressin, Eur. J. Pharmacol., 71 (1981) 33-42. 42 Unger, T., Becker, H., Petty, M., Demmert, G., Schneider, B., Ganten, D. and Lang, R.E., Differential effects of central angiotensin II and substance P on sympathetic nerve activity in conscious rats, Circ. Res., 56 (1985) 563-575. 43 Unger, T., Rohmeiss, E, Demmert, G., Ganten, D., Lang, R.E. and Luft, EC., Differential modulation of the baroreceptor reflex by brain and plasma vasopressin, Hypertension, Suppl. II, 8 (1986) 157-162. 44 Unger, T., Carolus, S., Demmert, G., Ganten, D., Lang, R.E., Maser-Gluth, C., Steinberg, H. and Veelken, R., Substance P induces a cardiovascular defense reaction in the rat: pharmacological characterization, Circ. Res., 63 (1988) 812-820. 45 Wormser, U., Laufer, R., Hart, Y., Chorev, M., Gilon, C. and Selinger, Z., Highly selective agonists for substance P receptor subtypes, EMBO J., 5 (1986) 2805-2808.
231
BRES 15873
Role of central tachykinin peptides in cardiovascular regulation in rats Yukio Takano 1, Akira Nagashima 1, Tetsuya Hagio 1, Kayoko Tateishi 2 and Hiro-o Kamiya 1 t Department of Pharmacology, Faculty of Pharmaceutical Sciences and 2The First Department of Biochemistry, School of Medicine, Fukuoka University, Fukuoka (Japan) (Accepted 20 March 1990) Key words: Substance P; Tachykinin peptide; Neurokinin B; Cardiovascular regulation; Vasopressin; Blood pressure
The mechanisms of action of tachykinin peptides thought to be involved in central cardiovascular regulation were examined. Intracerebroventricular injections (i.c.v.) of tachykinin peptides caused dose-dependent increases in blood pressure and heart rate. The pressor responses to substance P (SP) (10/~g, i.c.v.) and neurokinin A (NKA) (10/tg, i.c.v.) were blocked by peripheral administration of pentolinium or phentolamine, and partially attenuated by adrenalectomy. In contrast, the only initial pressor response to the neurokinin B (NKB) analogue senktide (10 pg, i.c.v.) was blocked by pentolinium or phentolamine. The pressor response to senktide was inhibited by pretreatment with a vasopressin V 1 receptor antagonist (d(CH2)5OMe(Tyr)AVP) (10/~g/kg, i.v.), and senktide (10 jug, i.c.v.) caused an increase in plasma vasopressin level. However, the vasopressin antagonist did not influence the SP- and NKA-induced pressor responses. These results suggest that central SP and NKA increase the blood pressure and heart rate via sympathetic nerve activity, whereas central NKB increases the blood pressure mainly via release of vasopressin from the hypothalamus. INTRODUC~JON Central neural circuits play important roles in cardiovascular control by regulating autonomic functions or vasopressin release from the hypothalamus. Recently, m a n y neuropeptides that seem to be involved in central blood pressure control have been discovered. These neuropeptides have become of interest as endogenous substances regulating blood pressure like neurotransmitters, such as catecholamine, acetylcholine, serotonin, histamine and y-aminobutyric acid ( G A B A ) . Although much information has been obtained on these peptides (for reviews, see refs. 8, 27, 29), their effects on the cardiovascular system are still not clearly understood. Substance P (SP) is a m e m b e r of the tachykinin family of bioactive peptides, which includes neurokinin A ( N K A ) , neurokinin B (NKB), neuropeptide K (NPK) and neuropeptide y (NPy) (for reviews, see refs. 6, 18, 22). SP, N K A , N P K and NPy are derived from the preprotachykinin A gene, whereas NKB is derived from the preprotachykinin B gene 15-17'25'26. From cardiovascular findings, SP has become of interest as a neuropeptide regulating the blood pressure in the ventral medulla 5" 13,14,20,33 nucleus tractus solitarii (NTS) 4'10-12'21'23'36-38 and spinal cord 9'34'35. Intracerebroventricular injection (i.c.v.) of SP increases the blood pressure. This pressor response to SP seems to be regulated by increase in sympathetic nerve activity, without participation of vaso-
pressin 7"39'41'42'44. Previously, we found that the NKB analogue senktide (i.c.v.) also causes increase in blood pressure, and that its action seemed to be mediated by vasopressin secreted from the hypothalamus, because blockade of peripheral vascular vasopressin receptors inhibited the pressor response 24. However, little is known about the mechanisms of central cardiovascular regulation induced by tachykinin peptides. In the present study, we examined the central mechanisms of action of tachykinin peptides thought to be involved in cardiovascular regulation. First, we studied the effects of sympathetic blockade and a vasopressin antagonist on tachykinin-induced central pressor responses. Then, we examined the effects of tachykinin peptides (i.c.v.) on plasma vasopressin levels. MATERIALS AND METHODS Animals Male Wistar rats (250-300 g) were purchased from Kyudo (Kumamoto, Japan). The animals were kept in a room at 20-25 °C with a 12 h light-dark cycle (light on at 07.00 h) and were given free access to commercial food and tap water. Cardiovascular response Experiments on cardiovascular responses were performed as described previously 23,24. Rats were anesthetized with urethane (1 g/kg i.p.). Then one catheter was inserted into the right femoral artery for recording the blood pressure and heart rate, and another into the left femoral vein for administration of drugs. The body temperature was maintained at 37-38 °C with a heating pad. The
Correspondence: Y. Takano, Department of Pharmacology, Faculty of Pharmaceutical Sciences, Fukuoka University, Fukuoka 814-01, Japan. 0006-8993/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)
232 systolic blood pressure was maintained above 99 mm Hg by infusion of 4% Ficoll 70 (Nacalai Chemicals, Japan) in saline solution. Each rat was placed in a stereotaxic apparatus. A guide cannula (21-gauge stainless steel) was inserted into the lateral brain ventricle. The adrenal glands were removed bilaterally from some animals. A stabilization period of 30 min was allowed before administration of drugs. Peptides were dissolved in artificial cerebrospinal fluid (ACSF; in raM): NaCI 128, KCI 3, CaCI2 1.2, MgCI2 0.8, NaH2PO 4 0.25, NaHCO3 21 and glucose 3.4; pH 7.4) and 10pl volumes of solutions were injected manually over a period of 1 min with a Hamilton microsyringe. Injection of ACSF (i.c.v.) did not cause any detectable change in blood pressure or heart rate. After each experiment, Malachite-green was injected to confirm the site of injection.
Plasma vasopressin level Rats were decapitated 5 or 20 min after i.c.v, injections, and
A
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Fig. 1 summarizes the effects of intracerebroventricular injections (i.c.v.) of various tachykinin peptides on the mean arterial blood pressure and heart rate. The central pressor responses to the tachykinin peptides were dose-dependent, reaching maxima 4-6 rain after injections of the peptides and then persisting for at least 40 rain.
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SP and NKA were obtained from the Peptide Institute (Osaka, Japan). [1-(fl-fl-Cyclopentamethylene propionic acid, 2-(O-methyl)tyrosine] ArgS-vasopressin (d(CH2)5OMe(Tyr)AVP), [Pro7]-NKB, (suc-[Asp6,Me-PheS]-substance P(6-11) (senktide), [(AspS'6,Me PheS]substance P(5-11) (NHz-senktide) were from Peninsula (California, U.S.A). NPy was a gift from Dr. J.E. Krause (Washington University, St. Louis, U.S.A.). Data are expressed as means + S.E.M.s. Statistical significance was determined by the two-tailed Student's t-test, or the Dunnett's multiple comparison test.
Cardiovascular effects of tachykinin peptides 6. c. v.)
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RESULTS
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blood samples were collected in chilled tubes containing 1.2 mg EDTA-2Na. The plasma was stored at -70 °C, until just before use. Plasma samples (200/A) were mixed with an equal volume of 0.1 N HCI and passed through a Sep-Pac Cls cartridge (Millipore Waters, MA). The Sep-Pac C18 cartridge was washed with 4% acetic acid and then adsorbed vasopressin was eluted with 1.5 ml of methanol, and dried under nitrogen gas. Radioimmunoassay (RIA) was performed with a vasopressin-RIA kit (Mitsubishi-Yuka, Tokyo, Japan). The antiserum showed less than 0.01% cross-reactivity with oxytocin. The sensitivity of the assay was 0.063 pg/tube.
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pressure a n d h e a r t rate induced by tacbykinin peptides (i.c.v.) at different doses, (A) substance P, (B) neurokinin A, and (C) senktide. Values are means _+ S.E.M.s for 6-10 rats. Fig. 1. T i m e course o f the responses in b l o o d
233 A
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To determine the involvement of tachykinin peptides in the sympathetic nervous system, we tested the effects of sympathetic blocking agents on the cardiovascular responses. The ganglionic blocker, pentolinium, was administered i.v. 10 min before injections of tachykinin peptides (i.c.v.). Pentolinium (10 mg/kg, i.v.) reduced the basal blood pressure. Pretreatment with pentolinium significantly inhibited the increases in blood pressure and heart rate elicited by SP and NKA. Although pentolinium blocked the earlier pressor response induced by the NKB analogue senktide, the later blood pressure after injection of senktide was increased under sympathetic ganglionic blockade (Fig. 2). The effects of peripheral a- and fl-adrenoceptor blockades are summarized in Table I. Administration of an a-adrenoceptor antagonist, phentolamine (10 mg/kg, i.v.), reduced the basal blood pressure, and slightly reduced the basal heart rate. Under peripheral aadrenoceptor blockade, the central pressor responses induced by SP and NKA were blocked. In contrast, pretreatment with phentolamine blocked only the initial pressor response induced by senktide (i.c.v.) (Table I). Administration of a fl-adrenoceptor antagonist, propranolol (1 mg/kg, i.v.), decreased in heart rate. Under peripheral fl-adrenoceptor blockade, the pressor response and tachycardiac action induced by SP (10 pg, i.c.v.) were inhibited. Pretreatment with propranolol inhibited tachycardia induced by NKA or senktide, but did not change the NKA-induced pressor response. Propranolol had no effect on the hypertensive effect of senktide (10/~g, i.c.v.) (Table I).
Effect of adrenalectomy on cardiovascular responses to tachykinin peptides (i.c. v. ) Bilateral adrenalectomy had no effect on the basal blood pressure or heart rate. Injection of SP or NKA increased the blood pressure and heart rate in both the sham-operated control group and the adrenalectomized group, but sustained pressor responses (20-40 min after i.c.v, injections) were significantly less in the adrenolectomized group than in the control group (Table II). Adrenalectomy did not block the senktide-induced pressor response and had no effect on the tachycardia induced by tachykinin peptides (i.c.v.) (Table II).
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234 TABLE I Effects of a- and fl-adrenoreceptor blocking agents, and vasopressin V1 antagonist (i.v.) on the cardiovascular responses to tachykinin peptides (i.c.v.) Data are mean _+S.E.M. n
Substance P (10/~g, i.c.v.) Saline 11 Phentolamine (10 mg/kg, i.v.) 8 Propranolol (1 mg/kg, i.v.) 8 Vl-antagonist (10 mg/kg, i.v.) 7 Neurokinin A (10/~g, i.c.v.) Saline 13 Phentolamine (10 mg/kg, i.v.) 6 Propranoloi (1 mg/kg, i,v.) 6 Vl-antagonist (10/~g/kg, i.v.) 6 Senktide (10/tg, i.c.v.) Saline 6 Phentolamine (10mg/kg, i.v.) 7 Propranoloi (1 mg/kg, i.v.) 10 Vl-antagonist (10~g/kg, i.v. ) 6
Basal MAP
A M A P (mm Hg) a
BasalHR
5 rain
20 min
88.4_+6.70 46.6 _+1.24** 97.1_+3.26 76.9_+5.99
+23.2+2.09 +4.25 + 1.24"* +12.6 _+2.26** +19.6_+1.51
+12.8-+2.08 +1.00 + 1.45"* +0.50 _+2.75** +13.0_+2.48
89.8_+5.09 49.7_+3.16"* 97.3_+4.74 77.1_+4.58
+18.5_+1.73 +7.67 _+0.76* +23.7 _+3.81 +17.3 _+4 . 5 0
90.2+ 8.81 46.6_+2.89** 98.7_+4.70 81.1_+3.11
+15.3_+2.14 +6.00_+1.36" +14.6_+2.52 +5.17 _+1.76"
AHR (beats~rain)a 5 min
20 rain
422.4+13.1 422.3 _+19.1 321.4 _+6.36** 423.7_+15.2
+17.3-+5.15 +26.6 _+8.52 +19.8 + 5.07 +31.7_+12.7
+34.0 +25.9 +1.63 +17.1
+12.3_+1.80 +5.17 _+1.90 + 13.7 + 3.89 +14.3 _+2.21
441.0_+11.0 388.3 _+23.2* 295.2 + 12.3"* 426.6 _+7.33
+43.9+6.30 + 14,4 _+4.37 +38.2 _+ 15.4 +31.6 _+ 10.4 + 14.8 + 4.03* +14.8 _+4.03 +27.3 _+9.13 +20.8 + 8.29
+4.33+4.00 +7.71_+1.51 +11.1_+2.51 -18.8 _+2.36**
452.2_+21.6 418.0_+9.31 323.9+7.25** 410.1 _+15.3
+16.7_+6.29 +16.1_+3.78 +3.80+4.75 +19.2 _+6.31
_+5.50 _+6.47 _+5.25** + 6.22
+14.7 -+ 11.1 +5.57 -+ 6.21 +6.60 + 4.99 + 15.7 _+6.38
*P < 0.05 and **P < 0.01 vs corresponding value for saline control by the Dunnett's test. aChanges in mean arterial blood pressure (MAP) and heart rate (HR) at 5 and 20 min after i.c.v, administration of the peptides. Effects o f vasopressin antagonist on cardiovascular responses to tachykinin peptides (i.c.v.) To d e t e r m i n e whether vasopressin receptors in the peripheral vasculature are involved in the pressor responses observed after injections (i.c.v.) of tachykinin peptides, we injected the vasopressin V 1 receptor antagonist (i.v.) 10 min before tachykinin peptides. The vasopressin antagonist (10/~g/kg, i.v.) did not cause any detectable changes in either the blood pressure or the heart rate. P r e t r e a t m e n t with the vasopressin antagonist completely inhibited the sustained hypertension 4 - 4 0 min after injection induced by senktide (10 ~g, i.c.v.), and
reduced the blood pressure to below the basal level after 20 rain (Table I). It also inhibited the hypertension induced by other NKB analogues, such as [Pro7]-NKB and NH2-senktide (data not shown). It did not, however, significantly affect the increases in blood pressure induced by SP and N K A (Table I). Effects o f tachykinin peptides (i. c. v.) on p l a s m a vasopressin level Fig. 3 shows the plasma vasopressin levels 5 and 20 min after injections (i.c.v.) of tachykinin peptides. Injection of senktide (10/~g, i.c.v.) caused a m a r k e d increase in the
TABLE II Effects of bilateral adrenalectomy on the cardiovascular responses to tachykinin peptides (i.c. v. ) Data are mean + S.E.M. n
BasalMAP
A M A P (mm Hg) a 5 rain
20 min
BasalHR 40 min
Substance P (10~g, i.c.v.) Sham 7 85.8+4.90 Adrenalectomy 8 82.3+4.98
+17.9_+3.43 +9.14_+5.00 +15,4_+2.74 -0.38_+2.20
Neurokinin A (10/~g, i.c.v.) Sham 8 84.3+5.87 Adrenalectomy 8 86.6+8.04
+24.9-+3.00 +19.9-+3.93 +18.1+3.90 428.0+11.0 +28.0_+4.17 -6.25_+7.64** -9.88_+8.55** 420.8_+21.3
Senktide (10/~g, i.c.v.) Sham 8 88.2+2.85 Adrenalectomy 8 84.3_+4.68
+16.9-+2.26 +3.44-+3.21 +12.1_+3.51 -1.11_+5.62
AHR (beats~rain)~ 5 rain
20 min
40 rain
+6.14_+4.66 425.4_+13.3 +41.7_+7.29 +31.0_+12.2 +23.6_+10.6 -8.13_+4.32" 415.9_+13.7 +36.0_+7,21 +14.8_+4.46 +12.8_+5.88 +29.3+6.01 +45.8+11.8
+25.9_+9.22 +21.3_+12.2 +21.6_+13.9 +28.8_+8.73
+1.33+2.30 436.7-+10.1 +17.0_+4.86 +8.67_+8.33 +8.78+4.16 +0.78_+3.82 430.7_+12.0 +31.2+8.57 -2,11_+8.32 +5.00_+13.0
*P < 0.05 and **P < 0.01 vs corresponding value for sham-operated control by the two-tailed Student's t-test. aChanges in mean arterial blood pressure (MAP) and heart rate (HR) at 5, 20 and 40 min after i.c.v, administration of the peptides.
235 Angiotensin II 10ug (ICV)
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iii
/
senktide 10ug
W'aloF
ACSF
60
50 m
~
40
n >
30
~
20
/
NKA 10ug
~ -10 L 012 ..... 4 6
n
10
15 '
20 '
2'5
30
35 '
40 '
115
20
215
30
;5
40
~ 30
3'5
40
30
35
40
20[
10
n- T=IO k 5 min
20 min
tug oi-
Time after injections (i.c.v.) d¢ P < 0.05
~ P
I I I
I
I
0124
Fig. 3. Plasma vasopressin levels 5 and 20 min after injections (i.c.v.) of tachykinin peptides. Values are means + S.E.M.s for 4 rats. *P < 0.05 and **P < 0.01 vs corresponding value for ACSF control by the Dunnetrs test.
I
I
I
6 81 0
I
TIME (rain)
* P<0.05 ** P
H
ACSF
2or
plasma vasopressin level after 20 min (ACSF 8.93 + 1.73 pg/ml vs senktide 56.37 + 8.38 pg/ml), but the other tachykinin peptides (i.c.v.), SP, NKA and NPy, had no effect on the plasma vasopressin level after 5 or 20 min.
tu t~ 101" o- E
oL
N | ~ -10 I-
. . . . . . 4 6 8 1o 012
1~5
2Zo
i 25
15
2~)
25
Effect of angiotensin H antagonist fi.c.v.) on senktideinduced pressor response
4o[
Angiotensin II in the brain stimulates the releases of vasopressin, oxytocin and adrenocorticotropic hormone (ACTH), and induces increase in blood pressure. To determine the contribution of the central angiotensin II system to the hypertension induced by senktide, we examined the effect of pretreatment with the angiotensin II receptor antagonist saralasin on the senktide-induced pressor response. Injection of saralasin (10 big, i.c.v.) increased the basal blood pressure, which returned to the basal level 10 min after the injection. Pretreatment with saralasin (i.c.v.) completely abolished the pressor response induced by angiotensin II (10/~g, i.c.v.) but did not influence that induced by senktide (Fig. 4).
2ok
DISCUSSION
There are several reports that the central pressor response to SP seems to be mediated by the sympathetic nervous system, without participation of vasopressin in conscious rats 41"42"44. However, there are few reports of studies on the central roles of other tachykinin peptides, probably partly because of the poor solubility of NKB in water and the absence of selective ligands for the tachykinin receptor subtype. Recently, selective NKB receptor ligands, senktide, NHE-senktide and [Pro7] NKB, have been developed 19'45. The present results
_2oL I I I
I
I
I
012 4 6 8 1~1
I
I
TIME (rain) Fig. 4. Effect o f the angJotensin II receptor antagonist saralasin on
the pressor responses after injections of (A) angiotensin II and (B) senktide. Values are means _+ S.E.M.s for 4 rats. *P < 0.05, **P < 0.01 and ***P < 0.005 vs corresponding value for ACSF control by the two-tailed Student's t-test.
PPT-A GENE
PPT-B GENE
I
I SubstanceP I Neurokinin A
~
I
4""
$(*)
+)
NeurokininB
+)
|
I (+)
Fig. 5. Proposed model of the dual mechanisms of central cardiovascular regulation induced by tachykinin peptides. NTS, nucleus tractus solitarii; AP, area postrema; PPT, preprotachykinin.
236 indicate a possible mechanism of central cardiovascular regulation by tachykinin peptides. Injection (i.c.v.) of the NKB-analogue senktide caused dose-dependent increase in blood pressure, and this response was inhibited by pretreatment (i.v.) with vasopressin V 1 receptor antagonist. In addition, senktide caused a marked increase in the plasma vasopressin level. Very recently Polidori et al. 28 reported similar increases of the plasma vasopressin level by NKB analogues. These findings suggest that the central cardiovascular effects of NKB analogues are mediated mainly by vasopressin. Presumably, treatment with an NKB analogue (i.c.v.) stimulates the release of vasopressin from the paraventricular nucleus (PVN) and/or supraoptic nucleus (SON) of the hypothalamus through the NKB receptor (NK-3 subtype) (Fig. 5), because we found previously the PVN and SON had higher contents of NKB-like immunoreactivity than other areas 24. Moreover high densities of [3H]NKB binding sites are present in these areas 2'3°. But the initial pressor responses to NKB analogues were inhibited by pretreatment (i.v.) with pentolinium or phentolamine, suggesting that the sympathetic nervous system initially contributes to the NKB analogue-induced pressor response. Injection of angiotensin II into the lateral brain ventricle (i.c.v.) is reported to cause an increase in blood pressure by stimulating release of vasopressin 31. However, this central angiotensin II system is not involved in the senktide-induced pressor response, because the angiotensin II receptor antagonist saralasin (i.c.v.) did not influence the senktide-induced pressor response (Fig. 4). It is unknown why the senktide-induced pressor response was reduced below the basal level after pretreatment with the vasopressin V 1 receptor antagonist (Table I). One possible explanation of this finding is that vasopressin released into the plasma by senktide (i.c.v.) stimulates not only the vascular vasopressin V 1 receptor but also the vasopressin receptor subtype in the area postrema (AP). Recently vasopressin was found to increase the sensitivity of the baroreceptor reflex through the vasopressin receptor subtype in the area postrema 1' 3,32,40,43 Therefore, under blockade of the vascular
vasopressin V 1 receptor, the vasopressin released by senktide may enhance the baroreceptor reflex inhibition of sympathetic nerve activity through the vasopressin receptor in the area postrema (Fig. 5). This idea is supported by the finding that the sustained blood pressure induced by senktide was enhanced under sympathetic ganglionic blockade (Fig. 2). The NKA-induced pressor response was reduced by blockade of the sympathetic nervous system, and NKA did not alter the plasma vasopressin level. These findings suggest that the pressor responses to NKA as well as SP are mediated by the sympathetic nervous system. Another tachykinin peptide derived from the preprotachykinin A gene, NP~, (i.c.v.), also increased the blood pressure without influencing the plasma vasopressin level (unpublished data). Since the increases in heart rate induced by SP and NKA are blocked by the peripheral administration of pentolinium and propranolol, the heart rate responses to SP and NKA also seem to be mediated by the sympathetic nervous system. Removal of the adrenal glands attenuated the sustained pressor responses induced by SP and NKA. These findings suggest that SP and NKA activate sympathetic nerves to the vasculature and adrenal catecholamine secretion. Thus the initial pressor responses to SP and NKA may be due to the sympathetically mediated vasoconstriction, and the sustained pressor responses to adrenal catecholamine secretion. Fig. 5 is a diagram summarizing our proposal on the central cardiovascular regulations by tachykinin peptides. We postulate that central SP and NKA, derived from the preprotachykinin A gene, increase the blood pressure and heart rate via stimulation of sympathetic nerve activity, whereas central NKB, derived from the preprotachykinin B gene, increases the blood pressure via release of vasopressin from the hypothalamus. Acknowledgements. This work was supported in part by a grant from the Suzuken Memorial Foundation. We are grateful to Dr. A.D. Loewy (Washington University, St. Louis, MO) for helpful discussion. We thank T. Akiyama, K. Hasegawa, and K. Takakura for technical assistance.
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4 5
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