Sympathetic and parasympathetic interaction in vascular and secretory control of salivary glands in anaesthetised dogs.

The present study was undertaken to examine sympathetic-parasympathetic interactions in the regulation of salivary gland function, with special reference to the possible role of the sympathetic cotransmitter neuropeptide Y (NPY). In dogs anaesthetised with pentobarbitone, electrical stimulation of the parasympathetic nerve to the submandibular gland evoked an increase in glandular blood flow and salivary secretion. Sympathetic nerve stimulation evoked a significant prolonged attenuation of vasodilator and secretory responses to subsequent parasympathetic stimulation. This attenuation was not significantly altered by alpha- and beta-adrenoceptor blockade. Systemic administration of the sympathetic cotransmitter, NPY, mimicked the effect of the sympathetic stimulation by significantly attenuating vasodilatation and salivary secretion. The NPY Y1 receptor agonist, [Leu31, Pro34]NPY and the specific NPY Y2 receptor agonist N-acetyl[Leu28, Leu31]NPY 24-36 both significantly attenuated the vasodilatation and salivary secretion evoked by stimulation of the parasympathetic nerve. The NPY Y1 receptor antagonist, GR231118 significantly antagonised the attenuation of vasodilatation caused by both sympathetic stimulation and the NPY Y1 receptor agonist. GR231118 also inhibited the pressor response of NPY. Intra-arterial injection of methacholine and stimulation of the parasympathetic nerve both caused local vasodilatation in the gland which was significantly attenuated by pretreatment with sympathetic stimulation or the NPY Y1 agonist. The NPY Y2-specific agonist did not attenuate methacholine-induced vasodilatation but did attenuate vasodilatation evoked by parasympathetic stimulation. The results indicate that NPY as a sympathetic cotransmitter may have a role in the regulation of vascular secretory function of salivary glands.

Cardiorespiratory responses to intracerebroventricular injection of neuropeptide Y in anaesthetised dogs.

Cardiovascular and respiratory effects of intracerebroventricular (icv) administration of neuropeptide Y (NPY) and separate, preferential agonists for NPY Y1 and Y2 receptors were observed in anaesthetised dogs. Central injections of NPY resulted in significant cardiac slowing and decreases in arterial pressure. These cardiovascular effects were blocked by central injection of the NPY Y1- preferring antagonist 1229U91. Central injection of NPY did not have a significant effect on ventilation, but the NPY Y1 antagonist 1229U91 administered alone caused a significant increase in ventilation. The NPY Y1-receptor agonist [Leu31Pro34] NPY significantly decreased ventilation while the NPY Y2 receptor agonist N-acetyl [Leu28Leu31] NPY 24--36 significantly increased it. A similar inverse relationship was seen with respect to blood pressure, with the NPY Y1-receptor agonist [Leu31Pro34] NPY significantly decreasing blood pressure, while the NPY Y2 receptor agonist N-acetyl [Leu28Leu31] NPY 24-36 significantly increased it. These findings suggest a role for NPY Y1 receptors in pathways mediating decreases in ventilation and blood pressure, and for NPY Y2 receptors in those mediating increased ventilation and blood pressure.

Activity of centrally truncated analogues of neuropeptide Y at Y1 and Y2 receptor subtypes in vivo.

Neuropeptide Y (NPY), a sympathetic cotransmitter, has both prejunctional and postjunctional actions in the cardiovascular system. In anaesthetized rats, the bioassay system used here, NPY attenuates cardiac vagal action (a prejunctional or Y2 action) and increases blood pressure (a postjunctional or Y1 action). Several NPY analogues were tested against NPY. In these, centrally located amino acid sequences of various lengths were removed, and replaced with simpler 'spacers'. As the parent NPY molecule is considered to exist in a U-shape, these central truncations were intended to shorten the depth of the U, while maintaining the integrity of its two ends. The centrally truncated NPY analogues examined here retain activity at both receptor subtypes in vivo. These findings indicate that the U-shape of the parent molecule probably exists to assist stability, but that receptor binding occurs through sequences closer to the termini.

A novel neuropeptide Y analog, N-acetyl [Leu28,Leu31]neuropeptide Y-(24-36), with functional specificity for the presynaptic (Y2) receptor.

We have carried out functional and in vitro studies on a novel analog of neuropeptide Y which shows selectivity for the prejunctional or neuropeptide Y Y2 receptor. In anaesthetised rats N-acetyl [Leu28,Leu31]neuropeptide Y-(24-36) attenuates cardiac vagal action (a prejunctional or neuropeptide Y Y2 action) and has no significant pressor effects (postjunctional or neuropeptide Y Y1 action). In the human neuroblastoma cell line (SMS-KAN) which expresses and endogenous Y2-like neuropeptide Y receptor, N-acetyl [Leu28,Leu31]neuropeptide Y-(24-36) competes with peptide YY for binding sites with an IC50 of 0.5 +/- 0.1 nM. In contrast in a fibroblast Chinese hamster ovary cell line which expresses the cloned human neuropeptide Y Y1 receptor and is used to study changes in cytosolic calcium evoked by (a neuropeptide Y Y1 effect), N-acetyl [Leu28,Leu31]neuropeptide Y-(24-36) showed no activity even at high concentrations. The steric structure for this novel compound has been determined using proton nuclear magnetic resonance (NMR) spectroscopy and it is consistent with the C-terminal end of published structures of neuropeptide Y. We suggest acetylation and amino acid substitutions stabilise the molecule and allow it to bind only to the neuropeptide Y Y2 receptor.

[Leu31, Pro34] NPY, a selective functional postjunctional agonist at neuropeptide-Y receptors in anaesthetised rats.

Neuropeptide Y (NPY), a sympathetic cotransmitter, has both prejunctional and postjunctional actions in the cardiovascular system. In the bioassay system used here NPY attenuates cardiac vagal action (an indicator of prejunctional or Y2 action) and increases blood pressure (an indicator of postjunctional or Y1 action). [Leu31,Pro34]NPY has little or no prejunctional activity i.e. does not attenuate cardiac vagal action, but increases blood pressure as effectively as NPY. [Leu31,Pro34]NPY can therefore be used in functional experiments to distinguish between the two types of NPY receptor.

Pre- and postjunctional actions of neuropeptide Y and related peptides.

The effects of neuropeptide Y (NPY) and related peptide fragments on blood pressure and vagal action at the heart were compared in the anaesthetized rat. A change in vagal action was taken as a measure of presynaptic activity and a change in blood pressure was taken as a measure of postsynaptic activity. NPY, NPY-(13-36), PYY-(13-36), des-Ser22-NPY-(13-36) and a stabilized 13-36 analogue of NPY (ANA NPY) all exerted pressor actions and attenuated vagal action at the heart. The maximum vagal inhibitory or presynaptic action in order of potency was NPY, ANA-NPY, PYY-(13-36) significantly greater than NPY-(13-36), des-Ser22-NPY-(13-36). The order of potency for the half time of this effect was NPY, ANA-NPY significantly longer than PYY-(13-36) and NPY-(13-36), which were significantly longer than des Ser22-NPY-(13-36). For the pressor or postsynaptic effects, NPY increased blood pressure significantly more and for a longer duration than all the 13-36 fragments, which were not demonstrably different in this respect. These results are consistent with the proposal that there are two populations of NPY receptors. The C-terminal flanking peptide of NPY (CPON) and desamido-NPY had no effect on either vagal action at the heart or on blood pressure.