Control of βAR- and N-methyl-D-aspartate (NMDA) Receptor-Dependent cAMP Dynamics in Hippocampal Neurons

Norepinephrine, a neuromodulator that activates β-adrenergic receptors (βARs), facilitates learning and memory as well as the induction of synaptic plasticity in the hippocampus. Several forms of long-term potentiation (LTP) at the Schaffer collateral CA1 synapse require stimulation of both βARs and N-methyl-D-aspartate receptors (NMDARs). To understand the mechanisms mediating the interactions between βAR and NMDAR signaling pathways, we combined FRET imaging of cAMP in hippocampal neuron cultures with spatial mechanistic modeling of signaling pathways in the CA1 pyramidal neuron. Previous work implied that cAMP is synergistically produced in the presence of the βAR agonist isoproterenol and intracellular calcium. In contrast, we show that when application of isoproterenol precedes application of NMDA by several minutes, as is typical of βAR-facilitated LTP experiments, the average amplitude of the cAMP response to NMDA is attenuated compared with the response to NMDA alone. Models simulations suggest that, although the negative feedback loop formed by cAMP, cAMP-dependent protein kinase (PKA), and type 4 phosphodiesterase may be involved in attenuating the cAMP response to NMDA, it is insufficient to explain the range of experimental observations. Instead, attenuation of the cAMP response requires mechanisms upstream of adenylyl cyclase. Our model demonstrates that Gs-to-Gi switching due to PKA phosphorylation of βARs as well as Gi inhibition of type 1 adenylyl cyclase may underlie the experimental observations. This suggests that signaling by β-adrenergic receptors depends on temporal pattern of stimulation, and that switching may represent a novel mechanism for recruiting kinases involved in synaptic plasticity and memory.

去甲肾上腺素（Norepinephrine）是一种可激活β肾上腺素能受体（β-adrenergic receptors, βARs）的神经调节剂，能够促进学习记忆以及海马突触可塑性的诱导。在谢弗侧支CA1突触（Schaffer collateral CA1 synapse）处，多种形式的长时程增强（long-term potentiation, LTP）需要同时激活βARs与N-甲基-D-天冬氨酸受体（N-methyl-D-aspartate receptors, NMDARs）。为阐明介导βAR与NMDAR信号通路相互作用的机制，我们将海马神经元培养物中的环腺苷酸（cAMP）FRET成像（FRET imaging）技术，与CA1锥体神经元（CA1 pyramidal neuron）信号通路的空间机制建模（spatial mechanistic modeling）相结合。既往研究表明，在βAR激动剂异丙肾上腺素（isoproterenol）与细胞内钙（intracellular calcium）共同存在时，cAMP会协同生成。与之相反，我们的研究显示，当异丙肾上腺素的施加先于NMDA数分钟——这是βAR介导的LTP实验的典型时序——相较于单独施加NMDA的情况，NMDA引发的cAMP响应平均振幅会被减弱。模型模拟结果表明，尽管由cAMP、环腺苷酸依赖性蛋白激酶（cAMP-dependent protein kinase, PKA）以及4型磷酸二酯酶（type 4 phosphodiesterase）构成的负反馈回路（negative feedback loop）可能参与了NMDA诱导cAMP响应的减弱过程，但该回路不足以解释全部实验观测结果。取而代之的是，cAMP响应的减弱需要腺苷酸环化酶（adenylyl cyclase）上游的机制参与。我们的模型证实，由βAR的PKA磷酸化所引发的Gs向Gi转换（Gs-to-Gi switching），以及Gi对1型腺苷酸环化酶（type 1 adenylyl cyclase）的抑制作用，或许是上述实验观测结果的潜在机制。这表明β肾上腺素能受体的信号传导依赖于刺激的时间模式，而这种受体转换可能代表了一种招募参与突触可塑性与记忆过程的激酶的新机制。