Takako Makita

Endothelins are vascular-derived axonal guidance cues for developing sympathetic neurons

During development, sympathetic neurons extend axons along a myriad of distinct trajectories, often consisting of arteries, to innervate one of a large variety of distinct final target tissues. Whether or not subsets of neurons within complex sympathetic ganglia are predetermined to innervate select end-organs is unknown. Here we demonstrate in mouse embryos that the endothelin family member Edn3 (ref. 1), acting through the endothelin receptor EdnrA (refs 2, 3), directs extension of axons of a subset of sympathetic neurons from the superior cervical ganglion to a preferred intermediate target, the external carotid artery, which serves as the gateway to select targets, including the salivary glands. These findings establish a previously unknown mechanism of axonal pathfinding involving vascular-derived endothelins, and have broad implications for endothelins as general mediators of axonal growth and guidance in the developing nervous system. Moreover, they suggest a model in which newborn sympathetic neurons distinguish and choose between distinct vascular trajectories to innervate their appropriate end organs.

Frequency of mononuclear diploid cardiomyocytes underlies natural variation in heart regeneration

Adult mammalian cardiomyocyte regeneration after injury is thought to be minimal. Mononuclear diploid cardiomyocytes (MNDCMs), a relatively small subpopulation in the adult heart, may account for the observed degree of regeneration, but this has not been tested. We surveyed 120 inbred mouse strains and found that the frequency of adult mononuclear cardiomyocytes was surprisingly variable (>7-fold). Cardiomyocyte proliferation and heart functional recovery after coronary artery ligation both correlated with pre-injury MNDCM content. Using genome-wide association, we identified Tnni3k as one gene that influences variation in this composition and demonstrated that Tnni3k knockout resulted in elevated MNDCM content and increased cardiomyocyte proliferation after injury. Reciprocally, overexpression of Tnni3k in zebrafish promoted cardiomyocyte polyploidization and compromised heart regeneration. Our results corroborate the relevance of MNDCMs in heart regeneration. Moreover, they imply that intrinsic heart regeneration is not limited nor uniform in all individuals, but rather is a variable trait influenced by multiple genes.

Venous endothelin guides sympathetic innervation of the developing mouse heart

The mechanisms responsible for establishing correct target innervation during organ development are largely unknown. Sympathetic nerves traverse or follow blood vessels to reach their end-organs, suggesting the existence of vascular guidance cues that direct axonal extension. The sinoatrial node and the ventricle of the heart receive sympathetic innervation from the stellate ganglia (STG). Here we show that STG axons follow veins, specifically the superior vena cavae and sinus venosus, to reach these targets. We find that the election of these routes is determined by venous endothelium-derived endothelin-1, acting through its specific receptor Ednra expressed within a subpopulation of STG neurons. Furthermore, we demonstrate that Edn1-Ednra signaling is essential for functional regulation of the heart by sympathetic nerves. Our findings present venous Edn1 as a sympathetic guidance cue, and show how axon guidance mechanisms are coordinated with end-organ morphogenesis.

Coronin-1 and Calcium Signaling Governs Sympathetic Final Target Innervation

Development of a functional peripheral nervous system requires axons to rapidly innervate and arborize into final target organs and then slow but not halt their growth to establish stable connections while keeping pace with organ growth. Here we examine the role of the NGF-TrkA effector protein, Coronin-1, on postganglionic sympathetic neuron final target innervation. In the absence of Coronin-1 we find that NGF-TrkA-PI3K signaling drives robust axon growth and branching in part by suppressing GSK3β. In contrast, the presence of Coronin-1 (wild-type neurons) suppresses but does not halt NGF-TrkA-dependent growth and branching. This relative suppression in axon growth behaviors is due to Coronin-1-dependent calcium release via PLC-γ1 signaling, which releases PI3K-dependent suppression of GSK3β. Finally, we demonstrate that Coro1a−/− mice display sympathetic axon overgrowth and overbranching phenotypes in the developing heart. Together with previous work demonstrating the Coronin-1 expression is NGF dependent, this work suggests that periods before and after NGF-TrkA-induced Coronin-1 expression (and likely other factors) defines two distinct axon growth states, which are critical for proper circuit formation in the sympathetic nervous system.

Extracardiac control of embryonic cardiomyocyte proliferation and ventricular wall expansion

AIMS The strategies that control formation of the ventricular wall during heart development are not well understood. In previous studies, we documented IGF2 as a major mitogenic signal that controls ventricular cardiomyocyte proliferation and chamber wall expansion. Our objective in this study was to define the tissue source of IGF2 in heart development and the upstream pathways that control its expression. METHODS AND RESULTS Using a number of mouse genetic tools, we confirm that the critical source of IGF2 is the epicardium. We find that epicardial Igf2 expression is controlled in a biphasic manner, first induced by erythropoietin and then regulated by oxygen and glucose with onset of placental function. Both processes are independently controlled by retinoic acid signalling. CONCLUSIONS Our results demonstrate that ventricular wall cardiomyocyte proliferation is subdivided into distinct regulatory phases. Each involves instructive cues that originate outside the heart and thereby act on the epicardium in an endocrine manner, a mode of regulation that is mostly unknown in embryogenesis.

Nerve Control of Blood Vessel Patterning

Patterning of embryonic blood vessels occurs in association with nerves. In this issue of Developmental Cell, Li et al. (2013) report that nerve-derived chemokine Cxcl12 (also known as SDF-1), acting through its receptor Cxcr4, initiates blood vessel remodeling along cutaneous nerve trajectories to establish the proper pattern of cutaneous arteries.

Placode lineage contributes to enteric sensory neurons in response to endothelin signaling

Introduction: The CREB-BDNF pathway has an important role in regulation of hippocampal neurogenesis and therapeutic mechanism of antidepressant. Recent studies have demonstrated the isoform-specific DNA hypemethylation in BDNF promoters in rodents exposed to early developmental stress. Interestingly, S-adenosyl methionine (SAM), the major methyl donor in mammalian brain, has been reported for the relief of symptoms of depression. SAM is known to accelerate the methylation cycle of various molecules important for receptor function and neurotransmitter production. However, whether SAM has the opposite effect against SSRI antidepressant on the DNA methylation of BDNF/CREB genes and hippocampal neurogenesis are unknown. Methods: The dose effect of SAM on CREB/BDNF gene expression was measured with real-time PCR in the cultured rat embryonic hippocampal neural stem cells (NSCs). Immunofluorescence was used to investigate the influence of SAM, SSRI fluoxetion and 5-HT1A receptor agonist 8-OH DPAT on NSC neurogenesis. DNA methylation of BDNF/CREB promoters was detected by the Sequenom MassARRAY platform. Results: The mRNA expression of CREB, BDNF exon-VI containing transcript and total BDNF was significantly decreased in hippocampal NSCs following 48 h administration of 200 M but not 50 M SAM. SAM (200 M, 48 h) also decrease the proliferation of hippocampal NSCs, while the neuronand astrocyte-oriented differentiation weren’t affected. The methylation level was significantly increased to 12.5% in the CpG site 23–24 of BDNF exon-VI promoter and to 3.8% in the CpG site 9 of CREB promoter by SAM (200 M 48 h). The effect of SAM on CREB/BDNF expression, DNA hypemethylation and hippocampal NSC proliferation was significantly reversed by 48 h administration of 1 M fluoxetion and 5 M 8-OH DPAT. Discussion: SAM might act as the methyl donor under the particular regimen to inhibit gene expression of CREB-BDNF pathway and hippocampal neurogenesis through methylation activation, which could be attenuated by SSRI drugs and 5-HT1A receptor stimulation.

Venous endothelin guides sympathetic innervation of the heart

Introduction: The CREB-BDNF pathway has an important role in regulation of hippocampal neurogenesis and therapeutic mechanism of antidepressant. Recent studies have demonstrated the isoform-specific DNA hypemethylation in BDNF promoters in rodents exposed to early developmental stress. Interestingly, S-adenosyl methionine (SAM), the major methyl donor in mammalian brain, has been reported for the relief of symptoms of depression. SAM is known to accelerate the methylation cycle of various molecules important for receptor function and neurotransmitter production. However, whether SAM has the opposite effect against SSRI antidepressant on the DNA methylation of BDNF/CREB genes and hippocampal neurogenesis are unknown. Methods: The dose effect of SAM on CREB/BDNF gene expression was measured with real-time PCR in the cultured rat embryonic hippocampal neural stem cells (NSCs). Immunofluorescence was used to investigate the influence of SAM, SSRI fluoxetion and 5-HT1A receptor agonist 8-OH DPAT on NSC neurogenesis. DNA methylation of BDNF/CREB promoters was detected by the Sequenom MassARRAY platform. Results: The mRNA expression of CREB, BDNF exon-VI containing transcript and total BDNF was significantly decreased in hippocampal NSCs following 48 h administration of 200 M but not 50 M SAM. SAM (200 M, 48 h) also decrease the proliferation of hippocampal NSCs, while the neuronand astrocyte-oriented differentiation weren’t affected. The methylation level was significantly increased to 12.5% in the CpG site 23–24 of BDNF exon-VI promoter and to 3.8% in the CpG site 9 of CREB promoter by SAM (200 M 48 h). The effect of SAM on CREB/BDNF expression, DNA hypemethylation and hippocampal NSC proliferation was significantly reversed by 48 h administration of 1 M fluoxetion and 5 M 8-OH DPAT. Discussion: SAM might act as the methyl donor under the particular regimen to inhibit gene expression of CREB-BDNF pathway and hippocampal neurogenesis through methylation activation, which could be attenuated by SSRI drugs and 5-HT1A receptor stimulation.