Increased potency of neuropeptide Y to antagonize alpha2-adrenoceptor function in the nucleus tractus solitarii of the spontaneously hypertensive rat.

The regulation by neuropeptide Y of alpha2-adrenoceptors in the nucleus tractus solitarii was evaluated in the adult normotensive Wistar Kyoto rat and the adult spontaneously hypertensive rat. The microinjection of a submaximal dose of l-noradrenaline (800 pmol in 50 nl) alone into the nucleus tractus solitarii produced a significant reduction in the mean arterial blood pressure in either strain. The threshold dose (1 pmol in 50 nl) of neuropeptide Y(1-36) for the vasodepressor response in the Wistar Kyoto rat was five times higher than that (0.2 pmol in 50 nl) in the spontaneously hypertensive rat. Furthermore, neuropeptide Y(1-36) at 0.2 pmol in 50 nl could significantly counteract the vasodepressor response to l-noradrenaline (800 pmol in 50 nl) in the spontaneously hypertensive rat, but not in the Wistar Kyoto rat, in which 1 pmol in 50 nl of neuropeptide Y(1-36) must be employed to counteract the vasodepressor response to l-noradrenaline (800 pmol in 50 nl), although the vasodepressor responses are of a similar magnitude. The in situ hybridization and quantitative receptor autoradiographical experiments showed that the alpha2A-adrenoceptor messenger RNA levels and the B(max) value of the alpha2-adrenoceptor agonist [3H]p-aminoclonidine binding sites measured in the nucleus tractus solitarii of the spontaneously hypertensive rat were substantially lower than those in the Wistar Kyoto rat. The quantitative receptor autoradiographical results were consistent with the cardiovascular results and showed that in the spontaneously hypertensive rat, neuropeptide Y(1-36) at 1 nM led to a significant increase in the K(d) value of [3H]p-aminoclonidine binding sites. In the Wistar Kyoto rat, neuropeptide Y(1-36) produced this effect only at 10 nM. The present study provides evidence for an increase of the potency of neuropeptide Y(1-36) to antagonistically modulate alpha2-adrenoceptors in the nucleus tractus solitarii of the spontaneously hypertensive rat. This enhanced antagonistic action may partly be related to a reduction in the number of alpha2A-adrenoceptors in the nucleus tractus solitarii of the spontaneously hypertensive rat, since a decrease has been observed in the alpha2A-adrenoceptor messenger RNA levels and the alpha2-adrenoceptor binding sites in the spontaneously hypertensive rat. This increased potency of neuropeptide Y(1-36) to antagonize alpha2-adrenoceptor function in the nucleus tractus solitarii of the spontaneously hypertensive rat may contribute to the development of high blood pressure in this hypertensive strain.

Evidence for a differential modulation of the alpha-2 adrenoceptors by angiotensin II in the nucleus tractus solitarii of the spontaneously hypertensive and the Wistar-Kyoto normotensive rats.

An interaction between angiotensin II (Ang II) receptors and alpha 2-adrenoceptors was evaluated in the nucleus tractus solitarii (NTS) of the normotensive Wistar-Kyoto rat (WKY) and of the spontaneously hypertensive rat (SHR) using quantitative receptor autoradiography and cardiovascular analysis. In the WKY rat, Ang II promoted a dose-dependent increase in the IC50 value of l-noradrenaline when competing for ([3H]p-aminoclonidine ([3H]PAC) binding sites, which reached a maximum of 400% with 10 nM of Ang II and was associated with a small decrease in the B0 value (20%). In the SHR Ang II (0.1 nM) had an opposite effect leading to a decrease in the IC50 value of about 57%, and no change was observed in the B0 value. Saturation analysis also showed that Ang II (0.1 nM) increased the KD value of [3H]PAC in the WKY strain but in contrast decreased the KD value of [3H]PAC in the SHR. The Bmax value was not significantly changed neither in the WKY rat nor in the SHR. The cardiovascular analysis showed that a threshold dose of Ang II (0.05 pmol) counteracted the vasodepressor effect produced by l-noradrenaline coinjected in the NTS of the WKY rat. No effect was observed in heart rate. In the SHR no counteraction of the l-noradrenaline-induced vasodepressor effect was found, and in contrast a slight increase of the vasodepressor effect associated with a significant increase in the bradycardiac response was observed. The results give evidence for an antagonistic Ang II/alpha 2 receptor interaction in the cardiovascular part of the NTS of the WKY rat as previously observed in the Sprague-Dawley rat. However, this interaction is altered in the SHR, so that in this strain the Ang II/alpha 2 receptor interaction enhances alpha 2 affinity and possibly alpha 2 receptor function. This opposite effect observed in the SHR may represent one compensatory mechanism to counteract the development of high blood pressure in the SHR.

Bradykinin modulation of alpha 2-adrenoceptors in the nucleus tractus solitarii of the rat. An in vitro autoradiographical study.

The existence of an interaction between bradykinin (Bk) receptors and the alpha 2-adrenoceptors were evaluated by means of quantitative receptor autoradiography in the nucleus tractus solitarii (NTS) of the rat. In competition experiments using L-noradrenaline (0.1 nM to 10 microM) against [3H]p-aminoclonidine ([3H]PAC) (10 nM) it was observed that Bk produced an increase in the IC50 value of L-noradrenaline in a concentration response manner, which reached a maximum of about 100% with 10 nM of the peptide associated with a small decrease in the B0 value (15%). In saturation experiments Bk promoted a significant increase in the KD value of [3H]PAC (60%) and a decrease in the Bmax value (36%). The specific Bk B2 receptor antagonist HOE-140 fully counteracted the effect of Bk on the alpha 2-adrenoceptors as analyzed by the competition experiments. Furthermore, des-Arg9-Bk, a Bk analog which exhibits selective agonist activity to the Bk B1 receptor subtype did not produce any effect on the alpha 2-adrenoceptors, suggesting that the Bk B2 receptor subtype may be mediating the Bk action on the alpha 2-adrenoceptors in the NTS. The effect of Bk (10 nM) was analyzed together with GTP (0.1 nM) in competition experiments and no change in the ability of L-noradrenaline to compete for [3H]PAC binding sites was observed in the presence of GTP, suggesting that the receptor interaction between the Bk B2 receptors and the alpha 2-adrenoceptors may be a G-protein dependent mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)

Antagonistic regulation of alpha 2-adrenoceptors by neuropeptide Y receptor subtypes in the nucleus tractus solitarii.

The modulation of alpha 2-adrenoceptors by neuropeptide Y Y1 and neuropeptide Y Y2 receptor subtypes has been studied in the nucleus tractus solitarii of the male rat. The autoradiographical experiments showed that neuropeptide Y-(1-36), neuropeptide Y-(13-36), a selective neuropeptide Y Y2 receptor agonist, and [Leu31,Pro34]neuropeptide Y, a selective neuropeptide Y Y1 receptor agonist, in the nanomolar range increased the Kd value of the [3H]p-aminoclonidine binding sites in the above rank order of potency without changing the Bmax values. In contrast, in the competition experiments, the neuropeptide Y Y1 and the neuropeptide Y Y2 receptor agonists decreased and increased, respectively, with the same potency the IC50 value of l-adrenaline and especially of clonidine for the alpha 2-adrenoceptor agonist binding sites associated with an increase and a decrease of the B0 value, respectively. Cardiovascular experiments showed that microinjections of clonidine into the nucleus tractus solitarii induced dose-dependent vasodepressor and bradycardiac responses. Threshold doses for vasodepressor effects of neuropeptide Y-(1-36) and of the neuropeptide Y Y1 receptor agonist and for vasopressor effects of the neuropeptide Y Y2 receptor agonist significantly counteracted the vasodepressor action elicited by an ED50 dose of clonidine in the nucleus tractus solitarii, the bradycardiac action of clonidine also being counteracted by the neuropeptide Y Y2 but not the neuropeptide Y Y1 receptor agonist. The present results give indications for the existence of an antagonistic modulation of high affinity alpha 2-adrenoceptors by the neuropeptide Y Y1 and neuropeptide Y Y2 receptor subtype in the nucleus tractus solitarii which may contribute to a reduction of alpha 2-adrenoceptor-mediated cardiovascular depression.

Evidence for an antagonistic angiotensin II/alpha 2-adrenoceptor interaction in the nucleus tractus solitarii.

Interactions between alpha 2-adrenoceptors and angiotensin II receptors were evaluated in the nucleus tractus solitarii of the rat by means of quantitative receptor autoradiography and cardiovascular analysis. In binding experiments using l-noradrenaline to compete for [3H]p-aminoclonidine binding sites, angiotensin II (1 nM) increased the IC50 value of l-noradrenaline by 50%. The angiotensin AT1 receptor antagonist, DUP753 (losartan), not only blocked this action but also decreased the IC50 value of l-noradrenaline. The modulatory effect of angiotensin II was also evaluated after addition of both DUP753 and PD123319, an angiotensin AT2 receptor antagonist, and counteraction of the reduction in the IC50 value of l-noradrenaline was observed. In saturation experiments angiotensin II increased the KD and Bmax values of [3H]p-aminoclonidine binding sites, compatible with possible uncoupling of the alpha 2-adrenoceptors. Cardiovascular analysis demonstrated that a threshold dose of angiotensin II (0.05 pmol) counteracted the vasodepressor effect produced by an ED50 dose of l-adrenaline, l-noradrenaline or clonidine coinjected in the nucleus tractus solitarii. DUP753 fully blocked this in vivo modulation of alpha 2-adrenoceptors by angiotensin II. These findings suggest the existence of an antagonistic angiotensin AT1/alpha 2-adrenoceptor interaction in the nucleus tractus solitarii. Therefore, it can be surmised that the activation of angiotensin II AT1 receptors may reduce the transduction of the alpha 2-adrenoceptors and thus the alpha 2-mediated vasodepressor responses.

Selective modulation of the NPY receptors of the Y2 subtype by alpha 2 receptors in the nucleus tractus solitarii of the rat. A cardiovascular and quantitative receptor autoradiographical analysis.

The modulation of neuropeptide Y (NPY) receptors by alpha 2 receptors in the nucleus tractus solitarii (Sol) of the rat was evaluated using quantitative receptor autoradiography and measurements of mean arterial blood pressure and heart rate. The receptor autoradiographical experiments showed that clonidine (10 nM), a selective alpha 2 receptor agonist, induced a 59% increase in the B0 value and a 47% decrease in the IC50 value of NPY(1-36) when competing for [125I]peptide YY ([125I]PYY)-binding sites in the presence of [Leu31, Pro34]NPY (100 nM), a selective NPY Y1 receptor agonist, to block the binding to NPY Y1 receptors. In contrast, when NPY(13-36) (300 nM), a selective NPY Y2 receptor agonist, was used to block the binding to NPY Y2 receptors, clonidine (1-30 nM) did not affect the B0 value and the IC50 value of NPY(1-36) when competing for [125I]PYY-binding sites, suggesting that the stimulation of alpha 2 receptors can selectively increase the affinity of NYP(1-36) for the NPY Y2 receptor. Microinjections of threshold doses of adrenaline or clonidine into the Sol not only counteracted the vasopressor action of a close to ED50 dose of coinjected NPY(13-36), but also changed the vasopressor and tachycardic response produced by NPY(13-36) into a vasodepressor and bradycardic response. However, threshold doses of adrenaline or of clonidine microinjected into the Sol did not modify the vasodepressor responses to a close to ED50 dose of NPY(1-36) or of [Leu31, Pro34]NPY.(ABSTRACT TRUNCATED AT 250 WORDS)

Coinjections of NPY(1-36) or [Leu31,Pro34]NPY with adrenaline in the nucleus tractus solitarius of the rat counteract the vasodepressor responses to adrenaline.

The cardiovascular effects of adrenaline microinjected alone or together with neuropeptide Y (NPY) receptor agonists into the nucleus tractus solitarius (Sol) of the anaesthetized rat have been investigated in order to evaluate NPY/adrenergic receptor interactions. In the dose range 0.05-20 nmol, adrenaline microinjected unilaterally into the Sol produced significant dose-related reductions in mean arterial blood pressure and heart rate. The vasodepressor action of a close to ED50 dose of adrenaline (0.5 nmol) was significantly counteracted by a threshold dose of NPY (1-36) (1 pmol) and of the NPY Y1 receptor agonist [Leu31,Pro34]NPY (2.5 pmol) microinjected into the Sol, but not by a threshold dose of NPY(13-36)(50 fmol), a selective Y2 receptor agonist. The present study provides evidence for an antagonistic NPY Y1/adrenergic receptor interaction in the Sol of the rat, involved in cardiovascular regulation.

Autoradiographic evidence for a bradykinin/angiotensin II receptor-receptor interaction in the rat brain.

Using angiotensin II (ANG II) to compete with (3-[125I]iodotyrosyl-4, Sar1, Ile8)ANG-II ([125I]Sar1, Ile8)ANG II) for its binding sites in the nucleus of the solitary tract (nTS) and the paraventricular hypothalamic nucleus (PV) bradykinin (10 nM) reduced the IC50 value (48 nM) of ANG II, an action blocked by the bradykinin B2 antagonist HOE-140 (100 nM). In contrast, when analysing the high-affinity site (Kd 3.1 nM) for [125I]ANG II in the nTS bradykinin (10 nM) increased the Kd value. Thus, a central bradykinin/ANG II receptor interaction may exist involving a differential regulation of the high- and low-affinity ANG II receptors in the nTS. This regulation by bradykinin of angiotensin receptors in the nTS may help to explain the central vasopressor effect of bradykinin.

Localization of central pressor action of bradykinin in medulla oblongata.

The intracerebroventricular injection of bradykinin produces an increase in arterial blood pressure. The site of action for this effect has been reported to be in the lateral septal area, in the hypothalamus, or in the ventral portion of the third ventricle. Bradykinin injected into the fourth cerebral ventricle of unanesthetized rats produced a pressor effect with a shorter latency and a larger maximal effect than when injected in the third or lateral ventricles. Bradykinin in the fourth ventricle was also 10 times more potent than in the third ventricle and 100 times more potent than in the lateral ventricle. No changes in blood pressure were observed when bradykinin was injected into the cerebellum or in the subarachnoid space of the ventral surface of the brain or of the medulla. Microinjections in the medulla oblongata showed that the pressor responses were obtained when bradykinin was injected in the nucleus tractus solitarius or in the dorsal spinal trigeminal tract. No effect was observed after injections were given into the ventral, ventral lateral medulla, or other medullary regions. The data suggest that bradykinin may play a regulatory role in the central control of blood pressure by stimulating sites that are near the dorsal and dorsal lateral surfaces of the medulla and accessible to kinins in cerebrospinal fluid and in the cerebral arterial circulation.

Tissue kallikrein-kinin system in the brain and the regulation of arterial pressure.

The intracerebral administration of kininase inhibitors produced an increase in arterial pressure of the SHR but not in normotensive animals. The SHR were several fold more sensitive to the pressor response elicited by the intracerebral injection of bradykinin, however SHR with low blood pressure showed decreased sensitivity to the bradykinin pressor effect. The results suggest that an endogenous kinin system plays a role in the central regulation of blood pressure of the SHR.

Kinin receptors of the central nervous system of spontaneously hypertensive rats related to the pressor response to bradykinin.

1. Kinin analogues bradykinin (BK), T-kinin, Met-Lys-BK, Lys-Lys-BK, Des-Arg9-BK with agonist activity and D-Arg0-Hyp3-Thi5,8-D-Phe7-BK (DAHTDBK) and Arg9-Leu8-BK with antagonist activity were injected into the posterior portion of the fourth cerebral ventricle of unanaesthetized rats implanted with permanent cannulae and arterial pressure was measured directly from the abdominal aorta. 2. The spontaneously hypertensive rats (SHR) were more sensitive than normotensive Wistar rats (NWR) to the pressor effect of BK and other kinin analogues. The SHR did not differ in sensitivity of the pressor response to centrally administered angiotensin II or endothelin-1. 3. Experiments with selective kinin agonists and antagonists revealed that in the SHR, as in the NWR, the receptors which mediated the central pressor response are of the BK2 subtype. 4. Measurements of the pressor activity of kinins with different degrees of susceptibility to degradation, as well as experiments with kininase inhibitors, enalaprilat and CPP-Ala-Ala-Phe-pAB, suggest that the kininase activity in the central nervous system of SHR is reduced in comparison to that of NWR. 5. The SHR also showed increased sensitivity to BK and Lys-Lys-BK, compared with the NWR, when the kinins were injected following the administration of a mixture of the kininase inhibitors, suggesting that mechanisms other than kininase activity may play a role in the increased sensitivity of the SHR to the central pressor action of kinins. 6. An in vivo characterization of the kinin receptors which mediate the central pressor response showed that the interaction with DAHTDBK was reversible and of competitive nature. The pA2 in vivo estimated for the kinin receptors of the SHR was 0.7 logarithm units larger than that obtained in the NWR. 7. The kinin receptors which mediate the central BK pressor effect in the SHR are of the BK2 subtype and are similar to receptors in the NWR. The increased sensitivity to kinins in the SHR may be explained, at least in part, by their decreased kininase activity. At present it is impossible to conclude whether the difference observed in the pA2 represents an increased affinity of the kinin receptors or can be attributed to differences amongst strains in the enzymatic degradation of the antagonist.