MEDICAL HYPOTHESES AND RESEARCH
VOL. 5, No. 1/2, July 2009
M. N. Ghayur and L. J. Janssen  Med. Hypotheses Res. 5: 47‒56.
Real-time Imaging of Ca(2+)-handling in Intact Renal
Glomeruli Using Confocal Microscopy
Muhammad Nabeel Ghayur* and Luke Jeffrey Janssen
Department of Medicine, McMaster University, St. Joseph’s Hospital, Hamilton, Ontario,
Abstract. Glomeruli are filtering units in the kidneys. Being multicellular and complex
in structure, many aspects of glomerular function are yet to be elucidated. Most studies use
glomerular cells in culture, which may exhibit altered physiology compared to native cells.
Confocal microscopy has opened new avenues in exploring in situ glomerular function and
physiology. In this report, we propose experimenting with glomerular cells in renal cortical
slices and isolated intact glomeruli for Ca(2+)-handling studies. Cortical slices (100 µm thick)
were obtained from mice while intact glomeruli were isolated from rats using the sieving
method. These were loaded with fluo-4 and then placed in a confocal microscope. Fluo-4 was
excited using a 488 nm photodiode laser and images were collected at 1 frame/sec. Changes
in average fluorescence intensity (AFI) were interpreted as changes in [Ca(2+)]i. AFI
increased to 37.1 ± 6.7% and 84.3 ± 20.9% with Ang II (0.01 and 0.1 µM respectively).
Norepinephrine (10 µM), arginine vasopressin (0.1 µM) and K+ (30 mM) also elevated AFI
by 26.5 ± 6.8%, 22.3 ± 1.0% and 39.8 ± 10.3% respectively in the glomerular cells. Likewise
in isolated glomeruli, Ang II (0.1-10 µM), K+ (30-90 mM) and endothelin-1 (0.01-1 µM), all
showed elevation in [Ca(2+)]i. These results give an impetus for future studies examining Ca
(2+)-handling by confocal microscopy in glomerular cells using renal cortical slices and
isolated intact glomeruli. The results support the utility of this system for study of
glomerular physiology and pharmacology.
* Correspondence: Dr. Muhammad Nabeel Ghayur, Department of Medicine,
McMaster University, St. Joseph's Hospital, Room L-314, 50 Charlton Avenue East,
Hamilton L8N4A6, Ontario, Canada. Tel: 1-905-5221155 x 34327. Fax: 1-905-5406510. E-