Neuropeptide Y is a prejunctional inhibitor of vagal but not sympathetic inotropic responses in guinea-pig isolated left atria

Abstract

1. The effects of NPY and related peptides were examined on basal contractile force and nerve-mediated inotropic responses to electrical field stimulation of the guinea-pig isolated left atrium. 2. Electrical field stimulus (EFS)-inotropic response curves were constructed by applying 1-64 trains of four field pulses (200 Hz, 0.1 ms duration, 100 V) across isolated left atria (paced at 4 Hz, 2 ms, 1-4 V) within the atrial refractory period. Curves were constructed in presence of vehicle, propranolol (1 microM) or atropine (1 microM) to determine appropriate stimulus conditions. 3. The effects of PYY (1-10,000 nM), NPY (0.01-10 microM), N-Ac-[Leu28,31]NPY(24-36) (N-A[L]NPY(24-36); 0.01-10 microM) and clonidine (0.1-1000 nM) were examined on the positive and negative inotropic responses to EFS (eight trains, four pulses per refractory period). 4. NPY-related peptides had no effect on basal force of contraction nor on the inotropic concentration-response curves to bethanechol or isoprenaline. All three peptides inhibited vagally-mediated negative inotropic responses; rank order of potency PYY>NPY> or =N-A[L]NPY(24-36) was consistent with an action at prejunctional Y2-receptors. Clonidine concentration-dependently inhibited sympathetic inotropic responses. However, PYY, NPY and N-A[L]NPY(24-36) failed to mediate any significant inhibition of the positive inotropic response to EFS. 5. These data demonstrate that NPY is an effective inhibitor of vagal but not sympathetically-mediated inotropic responses in the guinea-pig isolated left atria. This may suggest that endogenously co-released NPY is important in mediating cross talk between efferent components of the autonomic nervous system modulating cardiac contractility, acting overall to sustain positive inotropic responses.

Figure 1

Representative trace recordings of the inotropic response to eight trains electrical field stimulation (EFS; each train was four pulses per refractory period, see Methods) in four separate guinea-pig isolated left atria. EFS was passed across the atria as indicated by the upward and downward arrows. (a) Control (vehicle) inotropic response to EFS. Second calibration bar on (a) indicates slowing of the chart speed from 25 to 5 mm min⁻¹; (b) inotropic response to EFS in the presence of 1 μM propranolol; (c) inotropic response to EFS in the presence of 1 μM propranolol +1 μM clonidine; (d) inotropic response to EFS in the presence of 1 μM atropine. Insert shows a schematic representation of the analysis of the inotropic responses to EFS (see Methods). b is the baseline force of contraction immediately prior to EFS. V₁ is the vagally-mediated decrease in punctate force of contraction, calculated as the difference between b and the peak force of contraction at the end of EFS (V₂). The sympathetically-mediated positive inotropic response to EFS is measured as the difference between the maximum increase in the force of contraction following EFS (s) and b.

Figure 2

Electrical field stimulus-inotropic response curves in guinea-pig isolated left atria pretreated for 30 min with either vehicle (n=4), propranolol (n=4) or atropine (n=4). Top panel is the positive inotropic response to 1-64 trains EFS (see Methods). Bottom panel is the negative inotropic response to EFS. Error bars are ±s.e.mean.

Figure 3

Effect of PYY and clonidine on the sympathetic response to field stimulation (EFS) of guinea-pig isolated left atria in the presence of atropine (1 μM). Curves are the positive inotropic response to eight trains EFS (each train was four pulses per refractory period; see Methods) expressed as percentage increase in baseline force (see Methods) following cumulative additions of either clonidine (0.1–1000 nM; n=5) or PYY (0.01–10 μM; n=5). The positive inotropic response to EFS with time in control atria (n=5) is shown for comparison. Vertical error bars are ±s.e.mean. Horizontal error bars are ±1 s.e.mean on the average fitted IC₅₀ for clonidine. C2 is the second control inotropic response to EFS immediately prior to addition of the first concentration of peptide, clonidine or vehicle.

Figure 4

Effect of peptides on vagal response to field stimulation (EFS) of guinea-pig isolated left atria in the presence of propranolol (1 μM) and clonidine (1 μM). Curves are the effect of cumulative additions of either PYY (1–1000 nM; n=5), NPY (0.01–10 μM; n=5) or N-Ac-[Leu^28,31]NPY(24–36) (N-A[L]NPY(24–36); 0.01–10 μM; n=5) on the negative inotropic response to eight trains EFS expressed as percentage inhibition of baseline force (each train was four pulses per refractory period; see Methods). The negative inotropic response to EFS with time in control atria (n=5) is shown for comparison. C2 is the second control response to eight trains EFS expressed as a percentage inhibition of baseline force for each of the treatment groups. Vertical error bars are ±s.e.mean for each concentration of peptide. Horizontal error bars indicate ±1 s.e.mean of the average fitted IC₅₀ for each curve.