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Overexpression of SERCA2a alters endoplasmic reticulum Ca 2+ handling in WKY but not SHR cardiac sympathetic neurons. A role for altered intracellular Ca 2+ handling in hypertensive sympathoexcitation?
Author(s) -
Shanks Julia,
Herring Neil,
Li Dan,
Johnson Errin,
Paterson David J
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.1007.19
Subject(s) - serca , thapsigargin , endoplasmic reticulum , endocrinology , medicine , intracellular , calcium , calcium in biology , extracellular , chemistry , atpase , biochemistry , enzyme
Increased sympathetic neurotransmission is well established in both animal and clinical models of hypertension. Altered cardiomyocyte calcium (Ca 2+ ) handling has also been implicated in heart failure, leading to the development of gene therapies to upregulate sarcoendoplasmic reticulum calcium ATPase (SERCA) in cardiomyocytes from heart failure patients. Dysregulation in neuronal Ca 2+ handling has previously been reported within isolated neurons of the spontaneously hypertensive rat (SHR). We tested whether overexpressing SERCA2a in isolated cardiac stellate neurons from SHR and Wistar Kyoto rats (WKY) could modulate intracellular Ca 2+ transients. Isolated neurons were incubated with either Ad‐MCherry‐SERCA2A or Ad‐MCherry‐Empty (control) for 24 hours immediately post isolation. Ca 2+ imaging using fura‐2AM (2.5μM) was carried out three days post transfection. Intracellular Ca 2+ changes were measured using Fura‐2AM in response to KCl, caffeine, thapsigargin and FCCP to mobilise intracellular Ca 2+ stores. In isolated SHR neurons SERCA2a overexpression facilitated greater depolarisation induced Ca 2+ transients than age matched WKY (WKY, Empty: 0.45±0.05 au, n=17. SERCA: 0.66±0.09 au, n=13. SHR, Empty: 0.65±0.06 au, n=18. SERCA: 0.80±0.04 au, n=25. * P <0.05), but no change in ER load (SHR. Empty: 0.16±0.04 au, n=18. SERCA: 0.17±0.02 au, n=25). Whereas ER Ca 2+ load was significantly increased post SERCA2a overexpression in the WKY (WKY. Empty: 0.02±0.02 au, n=17. SERCA: 0.11±0.02 au, n=13). Transmission electron microscopy shows a greater abundance of ER in SHR neurons. SERCA2a overexpression resulted in increased mitochondrial Ca 2+ load in both SHR and WKY neurons (WKY, Empty: 0.08±0.006 au, n=32. SERCA: 0.11±0.01 au, n=33. SHR, Empty: 0.63±0.009 au, n=25. SERCA: 0.10±0.01 au, n=32) (** P <0.01). Indicating that altered neuronal Ca 2+ handling in the SHR may be due to an already present abnormal ER Ca 2+ load, with upregulation of SERCA2a further effecting mitochondrial Ca 2+ handling and potentially metabolic function in these neurons. The effect of SERCA2a on neurotransmitter release was measured using 3 H‐norepinephrine overflow from SD rat atria in response to right stellate ganglia stimulation. Upregulation of atrial SERCA2a resulted in greater neurotransmitter release in compared to control (Empty: 98.7±20.5 cpm, n=7. SERCA: 186.5±28.41 cpm, n=8. * P <0.05). These results demonstrate that SERCA2a upregulation in cardiac sympathetic neurons with broad spectrum promoters results in increased neurotransmission and increased loading of Ca 2+ into intracellular stores that trigger greater Ca 2+ transients on subsequent neuronal depolarisation. Support or Funding Information British Heart Foundation