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The Autonomic Nervous System Regulates the Heart Rate through cAMP-PKA Dependent and Independent Coupled-Clock Pacemaker Cell Mechanisms

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ABSTRACT

Sinoatrial nodal cells (SANCs) generate spontaneous action potentials (APs) that control the cardiac rate. The brain modulates SANC automaticity, via the autonomic nervous system, by stimulating membrane receptors that activate (adrenergic) or inactivate (cholinergic) adenylyl cyclase (AC). However, these opposing afferents are not simply additive. We showed that activation of adrenergic signaling increases AC-cAMP/PKA signaling, which mediates the increase in the SANC AP firing rate (i.e., positive chronotropic modulation). However, there is a limited understanding of the underlying internal pacemaker mechanisms involved in the crosstalk between cholinergic receptors and the decrease in the SANC AP firing rate (i.e., negative chronotropic modulation). We hypothesize that changes in AC-cAMP/PKA activity are crucial for mediating either decrease or increase in the AP firing rate and that the change in rate is due to both internal and membrane mechanisms. In cultured adult rabbit pacemaker cells infected with an adenovirus expressing the FRET sensor AKAR3, PKA activity and AP firing rate were tightly linked in response to either adrenergic receptor stimulation (by isoproterenol, ISO) or cholinergic stimulation (by carbachol, CCh). To identify the main molecular targets that mediate between PKA signaling and pacemaker function, we developed a mechanistic computational model. The model includes a description of autonomic-nervous receptors, post- translation signaling cascades, membrane molecules, and internal pacemaker mechanisms. Yielding results similar to those of the experiments, the model simulations faithfully reproduce the changes in AP firing rate in response to CCh or ISO or a combination of both (i.e., accentuated antagonism). Eliminating AC-cAMP-PKA signaling abolished the core effect of autonomic receptor stimulation on the AP firing rate. Specifically, disabling the phospholamban modulation of the SERCA activity resulted in a significantly reduced effect of CCh and a failure to increase the AP firing rate under ISO stimulation. Directly activating internal pacemaker mechanisms led to a similar extent of changes in the AP firing rate with respect to brain receptor stimulation. Thus, Ca2+ and cAMP/PKA-dependent phosphorylation limits the rate and magnitude of chronotropic changes in the spontaneous AP firing rate.

No MeSH data available.


Non additivity of adrenergic and cholinergic stimulation. (A) Simulating the response to 20 nM ISO. The AP firing rate increases by 13%. (B) Simulating the response to 80 nM of CCh. The AP firing rate decreases by 16%. (C) Simulating the response to 80 nM of CCh followed by the addition of 20 nM of ISO. The AP firing rate increases by 6%. (D) Simulating the response to 20 nM of ISO followed by 80 nM of CCh. The AP firing rate increases by 6%. These set of simulations show that the response to treatment by ISO and CCh is not equal to the simple addition of the individual ISO and CCh effect on the AP firing rate (13–16% = −3% ≠ 6%).
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Figure 10: Non additivity of adrenergic and cholinergic stimulation. (A) Simulating the response to 20 nM ISO. The AP firing rate increases by 13%. (B) Simulating the response to 80 nM of CCh. The AP firing rate decreases by 16%. (C) Simulating the response to 80 nM of CCh followed by the addition of 20 nM of ISO. The AP firing rate increases by 6%. (D) Simulating the response to 20 nM of ISO followed by 80 nM of CCh. The AP firing rate increases by 6%. These set of simulations show that the response to treatment by ISO and CCh is not equal to the simple addition of the individual ISO and CCh effect on the AP firing rate (13–16% = −3% ≠ 6%).

Mentions: We evaluated the model's ability to reproduce some known sympathovagal effects. The compensatory effects of ChR stimulation by CCh to β-AR stimulation by ISO are shown in Figure 9. The model predicted that 88 nM of CCh would compensate for 10 nM of ISO (thus, a ratio 10:88 for rabbit SANCs). Furthermore, the model predicted a change in this ratio in response to an increase in ISO (Figure 9B). That is, relatively less CCh is needed to compensate for increased ISO treatment. Next we evaluated whether the effect of combined CCh and ISO is additive (Figure 10). Application of 20 nM of ISO increased the AP firing rate by 13% compared to the baseline (Figure 10A), while application of 80 nM CCh reduced the AP firing rate by 16% compared to the baseline (Figure 10B). However, when CCh was applied before (Figure 10C) or after ISO (Figure 10D), the net effect of both drugs was a 6% increase in the AP firing rate (i.e., not additive). Finally, we tested how and whether rapid application of CCh has a phasic effect on the AP firing rate (Figure 11). Application of short CCh pulse (see Section Methods for further details) has a phasic effect on the AP firing rate. The phasic changes in pacemaker cycle length depend on the timing of the vagal stimulation during the AP cycle. The greatest effect on AP cycle (appeared on the following beat) was observed when CCh was applied during early DD (Figure 11, see S3).


The Autonomic Nervous System Regulates the Heart Rate through cAMP-PKA Dependent and Independent Coupled-Clock Pacemaker Cell Mechanisms
Non additivity of adrenergic and cholinergic stimulation. (A) Simulating the response to 20 nM ISO. The AP firing rate increases by 13%. (B) Simulating the response to 80 nM of CCh. The AP firing rate decreases by 16%. (C) Simulating the response to 80 nM of CCh followed by the addition of 20 nM of ISO. The AP firing rate increases by 6%. (D) Simulating the response to 20 nM of ISO followed by 80 nM of CCh. The AP firing rate increases by 6%. These set of simulations show that the response to treatment by ISO and CCh is not equal to the simple addition of the individual ISO and CCh effect on the AP firing rate (13–16% = −3% ≠ 6%).
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Figure 10: Non additivity of adrenergic and cholinergic stimulation. (A) Simulating the response to 20 nM ISO. The AP firing rate increases by 13%. (B) Simulating the response to 80 nM of CCh. The AP firing rate decreases by 16%. (C) Simulating the response to 80 nM of CCh followed by the addition of 20 nM of ISO. The AP firing rate increases by 6%. (D) Simulating the response to 20 nM of ISO followed by 80 nM of CCh. The AP firing rate increases by 6%. These set of simulations show that the response to treatment by ISO and CCh is not equal to the simple addition of the individual ISO and CCh effect on the AP firing rate (13–16% = −3% ≠ 6%).
Mentions: We evaluated the model's ability to reproduce some known sympathovagal effects. The compensatory effects of ChR stimulation by CCh to β-AR stimulation by ISO are shown in Figure 9. The model predicted that 88 nM of CCh would compensate for 10 nM of ISO (thus, a ratio 10:88 for rabbit SANCs). Furthermore, the model predicted a change in this ratio in response to an increase in ISO (Figure 9B). That is, relatively less CCh is needed to compensate for increased ISO treatment. Next we evaluated whether the effect of combined CCh and ISO is additive (Figure 10). Application of 20 nM of ISO increased the AP firing rate by 13% compared to the baseline (Figure 10A), while application of 80 nM CCh reduced the AP firing rate by 16% compared to the baseline (Figure 10B). However, when CCh was applied before (Figure 10C) or after ISO (Figure 10D), the net effect of both drugs was a 6% increase in the AP firing rate (i.e., not additive). Finally, we tested how and whether rapid application of CCh has a phasic effect on the AP firing rate (Figure 11). Application of short CCh pulse (see Section Methods for further details) has a phasic effect on the AP firing rate. The phasic changes in pacemaker cycle length depend on the timing of the vagal stimulation during the AP cycle. The greatest effect on AP cycle (appeared on the following beat) was observed when CCh was applied during early DD (Figure 11, see S3).

View Article: PubMed Central - PubMed

ABSTRACT

Sinoatrial nodal cells (SANCs) generate spontaneous action potentials (APs) that control the cardiac rate. The brain modulates SANC automaticity, via the autonomic nervous system, by stimulating membrane receptors that activate (adrenergic) or inactivate (cholinergic) adenylyl cyclase (AC). However, these opposing afferents are not simply additive. We showed that activation of adrenergic signaling increases AC-cAMP/PKA signaling, which mediates the increase in the SANC AP firing rate (i.e., positive chronotropic modulation). However, there is a limited understanding of the underlying internal pacemaker mechanisms involved in the crosstalk between cholinergic receptors and the decrease in the SANC AP firing rate (i.e., negative chronotropic modulation). We hypothesize that changes in AC-cAMP/PKA activity are crucial for mediating either decrease or increase in the AP firing rate and that the change in rate is due to both internal and membrane mechanisms. In cultured adult rabbit pacemaker cells infected with an adenovirus expressing the FRET sensor AKAR3, PKA activity and AP firing rate were tightly linked in response to either adrenergic receptor stimulation (by isoproterenol, ISO) or cholinergic stimulation (by carbachol, CCh). To identify the main molecular targets that mediate between PKA signaling and pacemaker function, we developed a mechanistic computational model. The model includes a description of autonomic-nervous receptors, post- translation signaling cascades, membrane molecules, and internal pacemaker mechanisms. Yielding results similar to those of the experiments, the model simulations faithfully reproduce the changes in AP firing rate in response to CCh or ISO or a combination of both (i.e., accentuated antagonism). Eliminating AC-cAMP-PKA signaling abolished the core effect of autonomic receptor stimulation on the AP firing rate. Specifically, disabling the phospholamban modulation of the SERCA activity resulted in a significantly reduced effect of CCh and a failure to increase the AP firing rate under ISO stimulation. Directly activating internal pacemaker mechanisms led to a similar extent of changes in the AP firing rate with respect to brain receptor stimulation. Thus, Ca2+ and cAMP/PKA-dependent phosphorylation limits the rate and magnitude of chronotropic changes in the spontaneous AP firing rate.

No MeSH data available.