Horackova M, Huang M H, Armour J A, Hopkins D A, Mapplebeck C
Department of Physiology and Biophysics, Faculty of Medicine, Dalhousie University, Halifax, Canada.
Cardiovasc Res. 1993 Jun;27(6):1101-8. doi: 10.1093/cvr/27.6.1101.
The aim was to develop long term primary cocultures of adult ventricular myocytes with autonomic neurones dissociated from stellate or intrinsic cardiac ganglia. This was to provide an experimental model for the investigation of the regulatory role of autonomic neurones with respect to cardiac myocyte function.
Ventricular myocytes and stellate and intrinsic cardiac neurones were enzymatically dissociated from adult male guinea pigs and plated together on 13 mm cover slips; the cultures were maintained in an incubator for two to 10 weeks. The electrical properties of cultured myocytes and neurones were investigated by means of a conventional microelectrode technique and the spontaneous contractile activity of the myocytes was recorded by a video system. The electrical, contractile, and pharmacological properties of myocyte-neuronal networks were investigated by superfusing the cultures with various neuromodulators and blockers.
The electrical properties of the cultured myocytes and neurones were similar to those reported in other in vitro studies. Innervated and non-innervated cardiomyocyte cultures responded differently, however, to various pharmacological interventions. Spontaneous contractions were attenuated by tetrodotoxin (4 x 10(-7) M), beta adrenergic blockade, and nicotinic blockade more in cocultures than in cardiac myocyte cultures alone. On the other hand, the beta agonist isoproterenol increased the spontaneous beating in both types of cocultures more than in myocytes alone. The effects of the muscarinic agonist bethanechol and the muscarinic blocker atropine were not significantly different in innervated and non-innervated cultures. Nicotine induced either an increase or decrease in contractile rates of both cocultures and did not affect non-innervated myocyte cultures.
Cocultures of myocytes and autonomic neurones alter the responses of myocytes relative to cultures without neurones. The results suggest that functional contacts can be established between adult myocytes and dissociated neurones in primary cultures. Contractile rates of such myocyte cultures are influenced by the presence of neurones. Neurones innervating these cultures were modified by beta adrenergic, muscarinic, and nicotinic agents.
旨在建立成年心室肌细胞与从星状神经节或心脏固有神经节分离出的自主神经元的长期原代共培养体系。这是为了提供一个实验模型，用于研究自主神经元对心肌细胞功能的调节作用。
从成年雄性豚鼠中酶解分离出心室肌细胞、星状神经节和心脏固有神经元，并将它们共同接种在13毫米的盖玻片上；培养物在培养箱中维持两到十周。通过传统微电极技术研究培养的肌细胞和神经元的电特性，并用视频系统记录肌细胞的自发收缩活动。通过用各种神经调节剂和阻滞剂灌注培养物，研究肌细胞 - 神经元网络的电、收缩和药理特性。
培养的肌细胞和神经元的电特性与其他体外研究报道的相似。然而，有神经支配和无神经支配的心肌细胞培养物对各种药理干预的反应不同。在共培养物中，河豚毒素（4×10⁻⁷ M）、β肾上腺素能阻断和烟碱阻断对自发收缩的抑制作用比单独的心肌细胞培养物中更明显。另一方面，β激动剂异丙肾上腺素在两种共培养物中比在单独的肌细胞中更能增加自发搏动。毒蕈碱激动剂氨甲酰甲胆碱和毒蕈碱阻滞剂阿托品在有神经支配和无神经支配的培养物中的作用没有显著差异。尼古丁可使两种共培养物的收缩率增加或降低，而对无神经支配的肌细胞培养物没有影响。
与没有神经元的培养物相比，肌细胞和自主神经元的共培养改变了肌细胞的反应。结果表明，在原代培养中，成年肌细胞与分离的神经元之间可以建立功能联系。这种肌细胞培养物的收缩率受神经元的存在影响。支配这些培养物的神经元受到β肾上腺素能、毒蕈碱能和烟碱能药物的调节。
