Nerve growth factor transfer from cardiomyocytes to innervating sympathetic neurons activates TrkA receptors at the neuro-cardiac junction.

Sympathetic neurons densely innervate the myocardium with non-random topology and establish structured contacts (i.e. neuro-cardiac junctions, NCJ) with cardiomyocytes, allowing synaptic intercellular communication. Establishment of heart innervation is regulated by molecular mediators released by myocardial cells. The mechanisms underlying maintenance of cardiac innervation in the fully developed heart, are, however, less clear. Notably, several cardiac diseases, primarily affecting cardiomyocytes, are associated with sympathetic denervation, supporting the hypothesis that retrograde 'cardiomyocyte-to-sympathetic neuron' communication is essential for heart cellular homeostasis. We aimed to determine whether cardiomyocytes provide nerve growth factor (NGF) to sympathetic neurons, and the role of the NCJ in supporting such retrograde neurotrophic signalling. Immunofluorescence on murine and human heart slices shows that NGF and its receptor, tropomyosin-receptor-kinase-A, accumulate, respectively, in the pre- and post-junctional sides of the NCJ. Confocal immunofluorescence, scanning ion conductance microscopy and molecular analyses, in co-cultures, demonstrate that cardiomyocytes feed NGF to sympathetic neurons, and that this mechanism requires a stable intercellular contact at the NCJ. Consistently, cardiac fibroblasts, devoid of NCJ, are unable to sustain SN viability. ELISA assay and competition binding experiments suggest that this depends on the NCJ being an insulated microenvironment, characterized by high [NGF]. In further support, real-time imaging of tropomyosin-receptor-kinase-A vesicle movements demonstrate that efficiency of neurotrophic signalling parallels the maturation of such structured intercellular contacts. Altogether, our results demonstrate the mechanisms which link sympathetic neuron survival to neurotrophin release by directly innervated cardiomyocytes, conceptualizing sympathetic neurons as cardiomyocyte-driven heart drivers. KEY POINTS: CMs are the cell source of nerve growth factor (NGF), required to sustain innervating cardiac SNs; NCJ is the place of the intimate liaison, between SNs and CMs, allowing on the one hand neurons to peremptorily control CM activity, and on the other, CMs to adequately sustain the contacting, ever-changing, neuronal actuators; alterations in NCJ integrity may compromise the efficiency of 'CM-to-SN' signalling, thus representing a potentially novel mechanism of sympathetic denervation in cardiac diseases.

Role of low-voltage-activated calcium current and extracellular calcium in controlling the firing pattern of developing CA1 pyramidal neurons.

The firing pattern of individual neurons is an important element for information processing and storing. During the first weeks of development, there is a transitional period during which CA1 pyramidal neurons display burst-spiking behavior in contrast to the adult regular-firing pattern. Spike after-depolarizations (ADPs) constitute a major factor underlying burst-spiking behavior. Using current-clamp recordings, we studied ADP waveforms and firing patterns in CA1 pyramidal neurons of Wistar rats from 9 to 19 postnatal days (P9-19). The percentage of burst-spiking neurons increased up to P16, in correlation with the emergence of an active component in the ADP. The application of low-voltage-activated (LVA) calcium channel blockers such as nickel or mibefradil suppressed the generation of the active ADP component and burst-spiking behavior. In agreement with the development of the ADP waveform and burst-spiking behavior, voltage-clamp experiments in dissociated pyramidal neurons showed an increase in the LVA calcium current in P16-19 vs P9-12. Finally, we found that a reduction of extracellular calcium levels decreases the percentage of burst-spiking cells due to a reduction in the active component of the ADP. We conclude that a major contribution of LVA calcium channels to ADP determines the bursting capability of CA1 pyramidal neurons during a transitional postnatal period in contrast to adulthood.