Heterogeneity of prejunctional NPY receptor-mediated inhibition of cardiac neurotransmission

Abstract

Neuropeptide Y (NPY) has been proposed as the candidate inhibitory peptide mediating interactions between sympathetic and vagal neurotransmission in several species, including man. Here, we have defined the NPY receptors involved in modulation of cardiac autonomic neurotransmission using receptor-selective agonists and antagonists in the rabbit and guinea-pig isolated right atria. In isolated atrial preparations, sympathetically-mediated tachycardia (ST; with atropine 1 microM) or vagally-mediated bradycardia (VB; with propranolol 0.1-1 microM) in response to electrical field stimulation (EFS, 1-4 pulses) were tested 0-30 min after incubation with single concentrations of vehicle, NPY (0.01-10 microM), the Y2 receptor agonist N-Acetyl-[Leu28,31]NPY(24-36) (termed N-A[L]NPY(24-36)) or the Y1 receptor agonist [Leu31,Pro34]NPY (LP). The effect of NPY on the concentration-chronotropic response curves to isoprenaline and bethanechol were also assessed. Guinea-pig atria: NPY and N-A[L]NPY(24-36) caused concentration-dependent inhibition of VB and ST to EFS. Both peptides caused maximal inhibition of VB and ST within 10 min incubation and this remained constant. LP caused a concentration-dependent, transient inhibition of ST which was antagonized by the Y1-receptor antagonist GR231118 (0.3 microM), with apparent competitive kinetics. Rabbit atria: NPY (1 or 10 microM) had no effect on VB at any time point, but both NPY and LP caused a transient (approximately 10 min) inhibition of sympathetic tachycardia. This inhibition could be prevented by 0.3 microM GR231118. N-A[L]NPY(24-36) had no effect on ST. NPY had no effect on the response to beta-adrenoceptor stimulation by isoprenaline nor muscarinic-receptor stimulation by bethanechol in either species. Thus, in the guinea-pig, NPY causes a stable inhibition of both VB and ST to EFS via Y2 receptors and transient inhibition of ST via Y1 receptors. In contrast in the rabbit, NPY has no effect on the cardiac vagus and prejunctional inhibition of ST is transient and mediated by a Y1-like receptor (rather than Y2). Therefore it would be surprising if NPY plays a functional role in modulation of cardiac neurotransmission in the rabbit.

Figure 1

Left panel shows the effects of 30 min incubation with vehicle (water, n=6) or increasing concentrations of NPY (0.01 μM, n=6; 0.1 μM, n=6; 1 μM, n=6) on the bradycardic responses to electrical field stimulation in guinea-pig isolated right atria pretreated with propranolol (1 μM). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for vehicle or each concentration of NPY. Error bars are s.e.mean.

Figure 2

Left panel shows the effects of 30 min incubation with vehicle (water, n=5) or increasing concentrations of the Y₂ receptor agonist N-Acetyl-[Leu^28,31]NPY(24–36) (termed N-A[L]NPY(24–36); 0.01 μM, n=5; 0.1 μM, n=5; 1 μM, n=5) on the bradycardic responses to electrical field stimulation in guinea-pig isolated right atria pretreated with propranolol (1 μM). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for vehicle or each concentration of N-A[L]NPY(24–36). Error bars are s.e.mean.

Figure 3

Left panel shows the effects of 30 min incubation with vehicle (water, n=6), the Y₁ receptor agonist [Leu³¹,Pro³⁴]NPY (LP-NPY), the Y₁ receptor antagonist GR231118 (0.3 μM) or [Leu³¹, Pro³⁴]NPY in the presence of the Y₁ receptor antagonist GR231118 (0.3 μM) on the bradycardic responses to electrical field stimulation in guinea-pig isolated right atria pretreated with propranolol (1 μM). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for vehicle or each treatment group. Error bars are s.e.mean. n=5 per group.

Figure 4

Left panel shows the effects of 30 min incubation with vehicle (water, n=6) or increasing concentrations of NPY (0.1 μM, n=6; 1 μM, n=6, 10 μM, n=6) on the tachycardic responses to electrical field stimulation in guinea-pig isolated right atria pretreated with atropine (1 μM). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for vehicle or each concentration of NPY. Error bars are s.e.mean.

Figure 5

Left panel shows the effects of 30 min incubation with vehicle (water, n=5) or increasing concentrations of the Y₂ receptor agonist N-Acetyl-[Leu^28,31]NPY(24–36) (termed N-A[L]NPY(24–36); 0.1 μM, n=5; 1 μM, n=5; 10 μM, n=5) on the tachycardic responses to electrical field stimulation in guinea-pig isolated right atria pretreated with atropine (1 μM). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for vehicle or each concentration of N-A[L]NPY(24–36). Error bars are s.e.mean.

Figure 6

Left panel shows the effects of 30 min incubation with vehicle (water, n=6), the Y₁ receptor agonist [Leu³¹,Pro³⁴]NPY (LP-NPY), the Y₁ receptor antagonist GR231118 (0.3 μM) or [Leu³¹,Pro³⁴]NPY in the presence of the Y₁ receptor antagonist GR231118 (0.3 μM) on the tachycardic responses to electrical field stimulation in guinea-pig isolated right atria pretreated with atropine (1 μM). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for vehicle or each treatment group. Error bars are s.e.mean. n=5 per group.

Figure 7

Left panel shows the bradycardic response to electrical field stimulation in rabbit isolated right atria, pretreated with propranolol (1 μM) before and 30 min after incubation with vehicle (water, n=6) or NPY (1 μM, n=6). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for single atria pretreated with either vehicle or NPY (1 or 10 μM). Error bars are s.e.mean.

Figure 8

Left panel shows the effects of 30 min incubation with vehicle (water, n=6), NPY (10 μM), n=5), the Y₁ receptor agonist [Leu³¹,Pro³⁴]NPY (10 μM, n=5) or NPY (10 μM) in the presence of the Y₁ receptor antagonist GR231118 (0.3 μM, n=5) on the tachycardic responses to electrical field stimulation in rabbit isolated right atrial pretreated with atropine (1 μM). Right panel shows the responses to four pulses of electrical field stimulation (expressed as a percentage of control) versus time for vehicle or each treatment group. Error bars are s.e.mean

Figure 9

Tachycardic responses to electrical field stimulation in rabbit isolated right atria pretreated with atropine (1 μM). The data shown are the control stimulus response curves for the data set in Figure 7, prior to addition of peptide. Curves are 30 min after pretreatment with either vehicle (water, 15 μl, n=5) or GR231118 (0.3 μM, n=5). Error bars are s.e.mean.