Effects of neuropeptide Y (NPY) on isolated guinea-pig heart

Neuropeptide Y (NPY) is present in nerve fibres throughout the mammalian heart. We have elucidated the effects of NPY on the isolated papillary muscle and heart (Langendorff) from the guinea-pig. The paced papillary muscle was studied with regard to duration of the action potential, peak force, maximum rate of force development, time to peak force, and time from peak force to half relaxation; all these parameters were identical whether or not NPY (5 X 10(-7) M) was present in the medium. When a stimulation with trains of pulses was superimposed, the paced papillary muscle exhibited enhanced contractions. This increase in contractility was not observed in the presence of the beta-adrenoceptor antagonist propranolol (10(-6) M) and was thus considered to be adrenergically mediated. The latter (adrenergic) response was markedly attenuated by NPY. Since NPY did not interfere with the response to exogenous noradrenaline (NA) it is suggested that the peptide exerts a pre-junctional inhibitory affect on adrenergic nerve-mediated positive inotropy. Neuropeptide Y did not influence the electrocardiogram from the spontaneously beating heart (Lagendorff), nor did the peptide modify the positive chronotropic effect of exogenously applied NA. In conclusion, the results indicate that NPY is without effect on the heart muscle proper but that the sympathetic terminals of the heart possess pre-junctional receptors for NPY (and/or related peptides) related to suppression of stimulated NA release.     

Reserpine-induced potentiation of the inhibitory action of neuropeptide Y on the rat vas deferens neurotransmission

The inhibitory action of neuropeptide Y (NPY) on the muscular activity of the prostatic end of the rat vas deferens elicited by transmural electrical stimulation was examined in control and in reserpinized rats. Pretreatment with 1 mg/kg reserpine for 48 h induced a 6-fold increase in NPY potency. Likewise, the potency of clonidine to inhibit the electrically induced muscular activity or noradrenaline to contract the ductus musculature was also potentiated. It is hypothesized that reserpine via a denervation super-sensitivity-like process increases the density of the NPY receptors. The functional significance of NPY in the motor activity of the vas deferens is discussed.     

Electroimmunoblotting of neuropeptide Y: application to the rat vas deferens

In this study we have optimized the electroimmunoblotting conditions for neuropeptide Y (NPY). NPY standards and samples extracted from the rat vas deferens were separated on urea-sodium dodecyl sulphate gels. Densitometric scanning of the Coomassie Blue-stained gels allow a semi-quantitative analysis of NPY in the range of approximately 10(-11) to 10(-8) mol of NPY. Electroimmunoblotting of NPY was also shown to be best achieved overnight at 4 degrees C and with NC membranes of 0.22 micron. Under these conditions NPY extracted from the vas deferens has been efficiently electroimmunoblotted. Higher molecular weight NPY-reactive peptides were also detected that may be related to proteolytic processing of the NPY precursor.     

Co-release of neuropeptide Y (NPY) and noradrenaline from the sympathetic nerve terminals supplying the rat vas deferens; influence of calcium and the stimulation intensity.

Epididymal (E) and prostatic (P) segments of the rat vas deferens were incubated with tritium-labeled noradrenaline (NA); upon transmural electrical stimulation for 20 or 60 s (70 V, 1 ms, 3-35 Hz), the outflow of immunoreactive neuropeptide Y (ir-NPY) and NA was detected in the superfusion media. Ir-NPY was detected only following trains of 35 Hz for 60 s in both E and P. In contrast, tritium was released in a graded fashion following trains of 3, 15 or 35 Hz stimulation for 60 s in E, whereas in P it reached a plateau at frequencies larger than 15 Hz. The outflow of tritium, under present conditions, was dependent on the duration of the stimuli, while the release of ir-NPY was only evoked with stimuli of 60 s duration. In the absence of external Ca2+, neurotransmission was blocked and co-release of ir-NPY and NA was prevented.     

Inhibitory effect of neuropeptide Y (NPY) on the in vitro activity of dopamine-beta-hydroxylase.

Neuropeptide Y (NPY) is found to be costored with norepinephrine (NE) in vesicles of the nerve terminals. An endogenous inhibitor of dopamine-beta-hydroxylase (DBH), the synthetic enzyme of NE, has been mentioned. In an attempt to clarify the effect of NPY on DBH activities, an in vitro assay is carried out using chromatographic analysis of NE formation from dopamine. NPY (20-80 pmol/ml) produced a dose-dependent depression of NE formation catalysed by the purified bovine adrenal DBH. Lineweaver-Burke plots (Km = 1.1 mM, Vmax = 10 pmol/min/mg protein) showed a non-competitive inhibition in NPY (30 pmol/ml, IC50)-treated samples. Moreover, failure of denatured NPY even at maximum concentration to influence the DBH activities suggested the essential of natural form of NPY. Participation of sulfhydryl compound seems also negligible, because N-ethylmaleimide did not overcome the effect of NPY. These results indicate that NPY has the ability to inhibit the catalytic action of DBH.     

Effect of nonachlazine on adrenergic neurotransmission in the rat vas deferens

The effect of a new antianginal preparation, nonachlazine, on adrenergic neurotransmission in the isolated rat vas deferens was studied by recording contractions of the duct in response to transmural stimulation of postganglionic sympathetic nerves by the electric current or by application of noradrenalin (NA) or BaCl₂. The effect of nonachlazine on the NA content was studied spectrofluorometrically and the ability of the preparation to block the uptake of exogenous NA by the tissues also was investigated. Nonachlazine was found to have a moderate sympatholytic action, which was combined with its spasmolytic effect. Nonachlazine also has a well marked ability to block the reaccumulation of NA in the tissues.     

Structure activity relations for inhibition of neurotransmission in rat vas deferens by adenosine

Adenosine inhibits the isometric contractions of the rat vas deferens in response to field stimulation in vitro by presynaptic inhibition of transmitter release. In the present study the structure activity relations for the inhibition of neurotramsmission in the rat vas deferens by adenosine were examined. Adenosine and adenosine-N¹-oxide were the most potent inhibitors studied. 6-Methylaminopurine riboside, 6-hydroxylaminopurine riboside, 5′-deoxyadenosine and 5′-nitroadenosine were slightly less potent inhibitors. The common structural requirements for activity include a primary or secondary amine function at C₆ of the purine ring with little tolerance for major steric changes or substitutions on the sugar moiety. None of the analogues studied prevented the presynaptic inhibitory action of adenosine.     

Effects of cervical sympathetic nerve stimulation and neuropeptide Y (NPY) on cranial blood flow in the cat

Effects of cervical sympathetic nerve stimulation (SNS) at 10 Hz and intravenous infusion of neuropeptide Y (NPY), 10 and 100 pmol x kg body wt-1 x min-1 for 5 min, on regional blood flow in the cat were investigated with radioactive microspheres. Sympathetic nerve stimulation caused significant reductions in blood flows in the facial tissues including the eye. Alpha-adrenoceptor blockade with phenoxybenzamine and combined beta- and alpha-adrenoceptor blockade with propranolol and phenoxybenzamine abolished the effects of sympathetic nerve stimulation in most facial tissues except in the tongue, upper eyelid and masseter muscle. In most cranial tissues, neuropeptide Y reduced regional blood flow and increased vascular resistance. No effect of neuropeptide Y on vascular resistance was observed in the choroid. In the present study, evidence for a non-adrenergic component in sympathetic vasoconstriction was found in the tongue, upper eyelid and masseter muscle but not in the majority of feline facial tissues. Neuropeptide Y was a potent vasoconstrictor in many cranial tissues, while in parts of the uvea, the effects of neuropeptide Y were less pronounced.     

Asymmetric distribution of purinergic and adrenergic neurotransmission cooperates in the motor activity along the rat vas deferens

The distribution of purinergic and adrenergic responses in the epididymal and prostatic segment of the rat vas deferens were studied in vitro. Prazosin antagonizes the twitch elicited by electrical stimulation mainly in the epididymal segment while alpha,beta-methyleneadenosine 5'-triphosphate (alpha,beta-mATP) preferentially inhibits the response of the prostatic segment. Using both prazosin plus alpha,beta-mATP, the response to field stimulation was completely inhibited. Concentration-response curves revealed that adrenergic compounds elicited a greater contraction in the epididymal portion than in the prostatic end of the ductus. Purinergic compounds caused a contraction of greater magnitude in the prostatic portion. The results suggest that adrenergic and purinergic mechanisms are asymmetrically distributed along the vas deferens reflecting a gradient of adrenergic and purinergic receptors along the ductus.     

Demonstration of a neuropeptide Y (NPY)-like immunoreactivity in the pigeon retina

The distribution of neuropeptide Y (NPY)-like immunoreactivity in the pigeon retina was investigated by fluorescence immunohistochemistry. NPY-positive cells were found in central and peripheral retina. NPY somata were located in the proximal portion of the inner nuclear layer and their processes directed to the inner plexiform layer where they ramified in 3 immunoreactive bands. NPY might play a role as a neurotransmitter or neuromodulator in the pigeon retina.     

Current spread in the smooth muscle of the guinea-pig vas deferens

1. The membrane polarization in response to intracellular stimulation and external stimulation, the junction potentials evoked by nerve and field stimulation and the spontaneous junction potentials were studied in the guinea-pig vas deferens.

2. The responses to intracellular stimulation differed from those to external stimulation applied through a large electrode in the following ways: short time constant of the electrotonic potential; linearity of current—voltage relation; all-or-none spike only in a small proportion of the cells; high critical firing level; short latency; weak tendency for repetitive activity during depolarization; and sharp spatial decay of the response.

3. The difference between intracellular and external stimulation could be explained by differences in current distribution in the tissue, if many muscle fibres were aggregated in functional bundles, with three-dimensional cell-to-cell connexions, so that the membrane near an intracellular stimulating electrode was shunted by a large area of surrounding membrane.

4. The time course of the junction potentials depended on the manner by which they were produced. The junction potential evoked by hypogastric nerve stimulation was recorded in every cell with almost the same amplitude. The spontaneous junction potential decayed very sharply with distance and the time course of the falling phase was about 10 times faster than that of the evoked junction potential.

The difference between the time course of the junction potentials was also explained by the difference in current distribution in the tissue.

     

New immunomodulatory role of neuropeptide Y (NPY) in Salmo salar leucocytes

Neuropeptide Y (NPY) plays different roles in mammals such as: regulate food intake, memory retention, cardiovascular functions, and anxiety. It has also been shown in the modulation of chemotaxis, T lymphocyte differentiation, and leukocyte migration. In fish, NPY expression and functions have been studied but its immunomodulatory role remains undescribed. This study confirmed the expression and synthesis of NPY in S. salar under inflammation, and validated a commercial antibody for NPY detection in teleost. Additionally, immunomodulatory effects of NPY were assayed in vitro and in vivo. Phagocytosis and superoxide anion production in leukocytes and SHK cells were induced under stimulation with a synthetic peptide. IL-8 mRNA was selectively and strongly induced in the spleen, head kidney, and isolated cells, after in vivo challenge with NPY. All together suggest that NPY is expressed in immune tissues and modulates the immune response in teleost fish.     

Opposite effects of centrally administered neuropeptide Y (NPY) on locomotor activity of spontaneously hypertensive (SH) and normal rats

Centrally administered neuropeptide Y has been shown to produce sedation manifested by a suppression of locomotor activity and a synchronizing effect on the EEG pattern in normal rats. It has been suggested that this sedative effect of NPY is largely due to a facilitation of the alpha 2-adrenergic transmission line. In the spontaneously hypertensive (SH) rat, the NPY-induced up-regulation of alpha 2-adrenoceptors observed in normal rats is absent, and NPY produces a desynchronization of the EEG. In the present study, we have therefore examined the effects of NPY on locomotor activity of SH rats and of inbred controls of the Wistar-Kyoto (WKy) strain, in both morning and evening sessions. In morning sessions, NPY (0.2-5.0 nmol intracerebroventricularly, i.c.v.) increased locomotor activity of SH rats in a dose-related manner. WKy rats were largely inactive per se, and no effects of NPY could be detected. In evening sessions, when spontaneous activity is high, NPY (I.0 nmol i.c.v.) still increased the activity of the SH rats. In WKy rats, an activity suppression similar to that previously reported for normal Sprague-Dawley rats was seen. The present results indicate that the sedative action of NPY in different rats strains correlates with the ability of the peptide to up-regulate alpha 2-adrenergic receptors.     

Influence of castration on the membrane reactivity of the guinea-pig vas deferens

The guinea-pig vas deferens is a quiescent muscle which after castration undergoes atrophy and shows spontaneous contractions preceded by membrane spike activity. The influence of castration on the spontaneous release of neurotransmitters and on the internal concentration of sodium and potassium ions was studied. Utilizing the microelectrode technique it was shown that castration induces a partial depolarization (10 mV) of the cell membrane, but did not change the frequency of spontaneous excitatory junction potentials (SEJPs) of guinea-pig vas deferens. However, the time-course and the amplitude of the SEJPs were increased after castration, probably because of changes in membrane properties related to organ atrophy. Castration probably promotes a change in the ionic permeability of the smooth muscle fibre, since the ratio pNa/pK was twice that of control muscles.     

A neuropeptide Y (NPY)-related peptide is present in the river lamprey CNS

Pancreatic polypeptide (PP)-like material from brain + spinal cord, and retina extracts of Lampetra fluviatilis was studied by HPLC and RIA. The brain + spinal cord extract showed a complex elution profile with multiple peaks of immunoreactivity. The retina extract showed a much simpler pattern with a single significant peak along with a trace of a second peak corresponding to the latest and penultimate peaks in the brain extract. Twenty-one out of 36 residues could be sequenced from the latest eluting peak in the brain extract. This sequence showed 81% identity with porcine neuropeptide (NPY) suggesting that both the brain/spinal cord and retina of the river lamprey contain a peptide homologous to NPY.