Functional Coupling with Cardiac Muscle Promotes Maturation of hPSC-Derived Sympathetic Neurons

Oh, Yohan; Cho, Gun-Sik; Li, Zhe; Hong, Ingie; Zhu, Renjun; Kim, Minjeong; Kim, Yong Jun; Tampakakis, Emmanouil; Tung, Leslie; Huganir, Richard L.; Dong, Xinzhong; Kwon, Chulan; Lee, Gabsang

doi:10.1016/j.stem.2016.05.002

Cited by 98 publications

(149 citation statements)

References 44 publications

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“…8d), and we did not detect noradrenaline release by HPLC ( data not shown ), suggesting an immature AN-like phenotype. A recent study showed that only upon co-culture with mouse cardiomyocytes, HES-derived sympathetic neurons express such late markers, further supporting our immaturity hypothesis 26 . When we applied the AN-like protocol to FD-PSC lines, we found that cells from severe FD patients could not effectively generate the CD49D + NC, in contrast to control and mild FD cells (Fig.…”

Section: Resultssupporting

confidence: 78%

Capturing the biology of disease severity in a PSC-based model of familial dysautonomia

Zeltner

Fattahi

Dubois

et al. 2016

Nat Med

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Familial Dysautonomia (FD) is a debilitating disorder that affects derivatives of the neural crest (NC). For unknown reasons, FD patients show marked differences in disease severity despite carrying the identical, homozygous IKBKAP point mutation. Here, we present disease related phenotypes in pluripotent stem cells (PSCs) that capture FD severity. Cells from severe but not mild patients exhibit impaired specification of NC derivatives including autonomic and sensory neurons. In contrast, both severe and mild FD cells show defects in peripheral neuron survival, indicating neurodegeneration as culprit for mild FD. While genetic repair of the FD mutation reversed early developmental NC defects, sensory neuron specification was not restored indicating that other factors may contribute to disease severity. Whole exome sequencing identified candidate modifier genes for severe FD. Our study demonstrates that PSC-based modeling is sensitive in recapitulating disease severity, which presents an important step on the road towards personalized medicine.

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Section: Resultssupporting

confidence: 78%

Capturing the biology of disease severity in a PSC-based model of familial dysautonomia

Zeltner

Fattahi

Dubois

et al. 2016

Nat Med

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“…More recently, we have investigated the biophysics of neuro‐cardiac signalling, aiming to address whether the neuronal effects on the heart were the result, in accordance with the archetypal description of cardiac physiology, of the unabridged propagation of sympathetic neurotransmitters throughout the myocardium, or whether neuro‐cardiac communication was confined to specific junctional sites, similar to those involved in skeletal muscle contraction (Hall & Sanes, ; Homan & Meriney, ). This latter hypothesis arose from the results of several previous studies obtained in vitro (Chun & Patterson, ; Shcherbakova et al ., ; Oh et al ., ) and ex vivo (Fukuda et al ., ), suggesting that specific sympathetic synapses may exist in the heart. By applying SN optogenetics, we demonstrated in vivo that neurotransmission underlying the rapid and efficient chronotropic effect of SN activation depended on the local release of NE at the intercellular contact site, with features typical of synaptic transmission (Prando et al ., ).…”

Section: Introductionmentioning

confidence: 98%

Cardiac sympathetic innervation network shapes the myocardium by locally controlling cardiomyocyte size through the cellular proteolytic machinery

Pianca

Bona

Lazzeri

et al. 2019

The Journal of Physiology

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Key points The heart is innervated by a dense sympathetic neuron network which, in the short term, controls chronotropy and inotropy and, in the long term, regulates cardiomyocyte size. Acute neurogenic control of heart rate is achieved locally through direct neuro‐cardiac coupling at specific junctional sites (neuro‐cardiac junctions). The ventricular sympathetic network topology is well‐defined and characteristic for each mammalian species. In the present study, we used cell size regulation to determine whether long‐term modulation of cardiac structure is achieved via direct sympatho‐cardiac coupling. Local density of cardiac innervation correlated with cell size throughout the myocardial walls in all mammalian species analysed, including humans. The data obtained suggest that constitutive neurogenic control of cardiomyocyte trophism occurs through direct intercellular signalling at neuro‐cardiac junctions. Abstract It is widely appreciated that sympathetic stimulation of the heart involves a sharp increase in beating rate and significant enhancement of contractility. We have previously shown that, in addition to these evident functions, sympathetic neurons (SNs) also provide trophic input to cardiomyocytes (CMs), regulating cell and organ size. More recently, we have demonstrated that cardiac neurons establish direct interactions with CMs, allowing neuro‐cardiac communication to occur locally, with a ‘quasi‐synaptic’ mechanism. Based on the evidence that cardiac SNs are unevenly distributed throughout the myocardial walls, we investigated the hypothesis that CM size distribution reflects the topology of neuronal density. In vitro analyses of SN/CM co‐cultures, ex vivo confocal and multiphoton imaging in clarified hearts, and biochemical and molecular approaches were employed, in both rodent and human heart biopsies. In line with the trophic effect of SNs, and with local neuro‐cardiac communication, CMs, directly contacted by SNs in co‐cultures, were larger than the non‐targeted ones. This property reflects the distribution of CM size throughout the ventricles of intact mouse heart, in which cells in the outer myocardial layers, which were contacted by more neuronal processes, were larger than those in the less innervated subendocardial region. Such differences disappeared upon genetic or pharmacological interference with the trophic SN/CM signalling axis. Remarkably, CM size followed the SN distribution pattern in other mammals, including humans. Our data suggest that both the acute and chronic influence of SNs on cardiac function and structure is enacted as a result of the establishment of specific intercellular neuro‐cardiac junctions.

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“…In 2015 [30], experiments demonstrated that having sympathetic neurons present in in-vitro cardiac cultures delays cardiomyocyte cell cycle withdrawal and transiently limits hypertrophy via a β-adrenergic signaling pathway, which suggests that sympathetic innervation can regulate cardiomyocyte numbers during the postnatal period. Developmental changes may occur in neurons as well: Oh et al 2016 reported increased maturation of hiPSC-derived sympathetic neurons in their cardiac neuron co-culture system [31]. Coppen et al have suggested the existence of post-natal changes in connexin expression in the developing fetal heart [32].…”

Section: Discussionmentioning

confidence: 99%

Optical interrogation of sympathetic neuronal effects on macroscopic cardiac monolayer dynamics

Burton

Tomek

Ambrosi

et al. 2019

Preprint

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Alterations in autonomic function are known to occur in cardiac conditions including sudden cardiac death. Cardiac stimulation via sympathetic neurons can potentially trigger arrhythmias. Dissecting direct neural-cardiac interactions at the cellular level is technically challenging and understudied due to the lack of experimental model systems and methodologies. Here we demonstrate the utility of optical interrogation of sympathetic neurons and their effects on macroscopic cardiac monolayer dynamics to address research targets such as the effects of adrenergic stimulation via the release of neurotransmitters, the effect of neuronal numbers on cardiac wave behaviour and the applicability of optogenetics in mechanistic in vitro studies. We combine photo-uncaging or optogenetic neural stimulation with imaging of cardiac monolayers to measure electrical activity in an automated fashion, illustrating the power and high throughput capability of such interrogations. The methods described highlight the challenges and benefits of co-cultures as experimental model systems. RESULTS:1] Stellate Sympathetic Neurons make contacts with cardiomyocytes: Scanning electron microscopy of sympathetic neurons growing in co-culture with cardiomyocytes in vitro shows connections between neurite extension and cardiac syncytium (Figure 1 A). The neuron bodies and extensions clearly make physical contact with myocytes (Figure 1 B, C, D, E, F, G, H, I, J). Immuno staining of co-cultures shows co-localisation of Synpasin and Beta2 receptors (Figure 1 K), sympathetic neurons show positive staining for Th (Figure 1 L), and fibronectin contamination in the co-cultures was assessed by staining with Vimentin (Figure 1 M), which showed low abundance. 2] Pattern formation is affected by the presence of neurons: Using dye-free imaging (Figure 2 A, B, C), we investigated how neuronal activation modulates cardiac patterns of activation in monolayer culture. We chose experimental conditions that spontaneously yield a wide variety of wavefront topologies within the imaging system's 16 x 16 mm field of view. Introduction of an additional cell type can potentially introduce heterogeneities that would impact wave front stability. Surprisingly, co-cultures displayed fewer wave breaks than their monoculture counterparts at similar plating densities (Figure 2 D). We broadly classified wave dynamics as simple (periodic target waves and single spiral wave reentry, Figure 2 D top row), or complex (single dominant spirals with additional irregular waves and multiple equally sized wavelets, Figure 2 D bottom row). While monocultures frequently displayed complex dynamics, co-cultures rarely displayed this behavior (Figure 2 E, P<0.05, chi-square test). Indeed, we observed wavelet reentry in only one of the co-culture preparations (6 isolations, 20 preparations). 3] Conduction velocity in spontaneously active co-cultures is faster than myocyte monocultures:We measured conduction velocity in unstimulated co-cultures (n=19) and myocyte monocultures (n=9). Figure 2 F s...

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Functional Coupling with Cardiac Muscle Promotes Maturation of hPSC-Derived Sympathetic Neurons

Cited by 98 publications

References 44 publications

Capturing the biology of disease severity in a PSC-based model of familial dysautonomia

Capturing the biology of disease severity in a PSC-based model of familial dysautonomia

Cardiac sympathetic innervation network shapes the myocardium by locally controlling cardiomyocyte size through the cellular proteolytic machinery

Optical interrogation of sympathetic neuronal effects on macroscopic cardiac monolayer dynamics

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