Swelling properties of mitochondrial preparations from the retina

Abstract

Retinae obtained from the eyes of cattle ( Bos taurus) immediately after slaughter have been separated into sub-cellular fractions by flotation and differential centrifugation. Outer segments of the rods were obtained by flotation into 30% sucrose. Other sub-cellular fractions were obtained by homogenizing the retinae freed of outer segments in 0.25 M sucrose and subjecting the homogenate to the following increasing sedimentation fields: 6 × 10 3 , 4.8 × 10 4 , 1.45 × 10 5 and 2.1 × 10 6 g/min respectively. Ribonucleic acid, deoxyribonucleic acid and protein contents, succinic-neotetrazolium reductase and cytochrome oxidase activities were determined in these fractions. These assays indicated that distinctive sub-cellular fractions had been isolated so that a fraction rich in mitochondria could be obtained. By measuring the changes in the optical density at 520 mμ of a mitochondrial suspension the osmotic behaviour of the mitochondria was examined in different concentrations of potassium chloride. The mitochondria were observed to behave as osmotic vesicles. Mitochondria in isotonic media were observed to swell when digitonin (10 −4 M) or thyroxine ( 2 × 10 −5 M ) was added. Other substances, such as calcium and phosphate ions, vitamin A alcohol and aldehyde, p-chloromercuribenzoate and formaldehyde appeared to have no or slight activity in this respect. Mitochondrial swelling caused by thyroxine or hypotonicity was reversed by the addition of a mixture of adenosine triphosphate (0.5 mM) Mg ++ (2 mM) and ox plasma albumin (0.66 mg/ml).