One of the most fundamental properties of biological membranes is that they are barriers to the permeation of polar molecules. This is largely due to the hydrophobicity of the membrane interior. However, this barrier property cannot be automatically extended to the permeation of small solutes such as oxygen, even though some experimental data suggest this to be true [1
]. The overall conclusion from recent studies of oxygen transport across model and cell plasma membranes is that membranes are not barriers to oxygen transport into the cell and mitochondrion, and created oxygen concentration differences across these membranes at physiological conditions are negligible (see Review [8
] and references therein). Only in membranes that contain a high cholesterol concentration and are dense with integral membrane proteins are lipids packed so closely that the solubility and diffusion of oxygen are severely reduced [9
]. These features are characteristic of the fiber cell plasma membrane of the eye lens, which shows extremely high cholesterol content [13
] and a high protein-to-lipid ratio [19
]. These characteristics suggest that the fiber cell plasma membrane can be a moderate barrier to oxygen transport. Additionally, oxygen, on its way to the center of the lens, has to cross many thousands of fiber cell plasma membranes; even a very small oxygen concentration difference across one membrane can, as a result, significantly contribute to the oxygen concentration gradient across the eye lens.
To prevent excessive light scattering and compromised lens transparency, fiber cells lose all subcellular organelles (including mitochondria) during maturation. In fact, the plasma membrane becomes essentially the only supramolecular structure of the matured fiber cells [23
]. The plasma membrane of the fiber cell has unique biochemical characteristics, with an extremely high cholesterol level [13
] and a high sphingomyelin content [15
], but only traces of polyunsaturated fatty acids [24
]. This composition ensures a physical (rigidity [26
]) and a chemical (resistance to peroxidation [28
]) membrane stability. It also allows us to infer that the fiber cell plasma membrane can be a moderate barrier to oxygen transport.
New fiber cells are continuously produced from the epithelial monolayer, and, as a result the adult lens contains two kinds of fiber cells: those located in the lens core, which are maturate and do not contain organelles, and those located in the lens outer layers, which are not yet maturate and contain organelles (including mitochondria). Mitochondrial respiration accounts for approximately 90% of oxygen consumption by the lens [29
], which suggests that the outer layers of the fiber cells can be responsible for low oxygen concentration in the lens nucleus.
It is important to note that oxygen concentration in the lens is very low, reaching a value close to zero in the lens core [30
]. It also has to be mentioned that already at the lens surface the oxygen concentration is low; values of oxygen partial pressure from 2–3 mmHg to 38 mmHg are reported in several vertebrate species [32
]. An increase in oxygen concentration is thought to be responsible for cataract formation following hyperbaric oxygen treatment or vitrectomy [39
]. Thus, the knowledge of oxygen concentration and distribution within the lens is of considerable interest. Few polarographic and optode measurements report oxygen partial pressure from 1 mmHg in the cat lens posterior cortex and nucleus [43
] to 10–22 mmHg in the rabbit lens [37
] and 0.8–4.0 mmHg in the human anterior cortex [44
]. These data indicate that to protect against age-related nuclear cataract formation, oxygen concentration in the lens has to be maintained at a very low level.
Because oxygen is constantly consumed, and oxygen consumption reactions are located inside the eye lens [29
], it follows that there must be a gradient in oxygen concentration across the fiber cell layers that build the eye lens. Oxygen consumption is necessary to maintain the low oxygen concentration inside the eye lens; otherwise, the concentration of oxygen should be equal to that outside the lens. The value of the oxygen concentration difference across each fiber cell layer is determined by the rate of oxygen consumption by cells confined inside this concentric fiber cell layer and the oxygen permeability coefficient of the cell layer, which is the sum of the permeability coefficient across a cytoplasm and across two plasma membranes. Profiles of the oxygen partial pressure across the isolated bovine lens were reported in an excellent paper by McNulty et al. [29
]. They assume, based on the value of the oxygen diffusion coefficient in plasma membrane from erythrocytes [45
], that oxygen crosses cell membranes readily, and that membranes provide essentially no resistance to the diffusion of oxygen, as the membranes are very thin barriers. We would like to contribute to this discussion by providing the values of the oxygen permeability coefficient across the membrane made of the total lipid extract from the plasma membrane of calf (bovine) lens and by evaluating the possible contribution of the fiber cell plasma membrane to the created gradients of oxygen concentration across the eye lens and to the process of maintaining the oxygen concentration in the eye nucleus at a very low level.
The oxygen permeability coefficient across the membrane can, in principle, be measured directly using stop-flow rapid-mixing apparatus to create an oxygen concentration difference across the membrane. These studies have been criticized because the presence of a thick (~2 μm), unmixed water layer on the membrane surface prevents immediate contact of the oxygenated solution with the membrane [46
]. In other studies, the oxygen permeability coefficient was calculated using the average (single) value of the oxygen diffusion coefficient and the average (single) value of the oxygen membrane concentration [45
]. It was customary to assume that oxygen dissolves and diffuses in the lipid bilayer in a similar way as in bulk hydrocarbon solvents. For example, the value of oxygen concentration in olive oil obtained by Battino et al. [51
] was used to evaluate oxygen transport within and across the lipid bilayer. Similarly, in the oxygen diffusion-consumption model, which describes the profile of the oxygen concentration across the bovine eye lens [29
], the average (single) value of the oxygen diffusion coefficient in membranes was used. Direct support for these assumptions, however, is lacking. Moreover, a number of reports have indicated that the oxygen diffusion-concentration product changes significantly from the membrane surface to the membrane center [9
]. Subczynski et al. [9
] developed the method for the calculation of the oxygen permeability coefficients across the membrane based on the profiles of the oxygen transport parameter (oxygen diffusion-concentration product) across the lipid bilayer, which can be obtained for membranes equilibrated with nitrogen and an air/nitrogen mixture without the need for creating fast-decaying oxygen concentration gradients. Our earlier papers [9
] give a solid base for this procedure.
In the present studies, we use our published earlier data of oxygen transport parameters in the lipid bilayer membrane made of the total lipid extract from the calf lens fiber cells [27
] to evaluate the oxygen permeability coefficient across this membrane. To elucidate better the major factors that determine membrane resistance to oxygen transport within and across the lens lipid membrane, we compared results obtained for this membrane with those obtained for membranes made of the equimolar 1-palmitoyl-2-oleoylphosphatidylcholine/cholesterol (POPC/Chol) mixture and of the pure POPC. The spin label oximetry measurement splits the lipid bilayer membrane into three well distinguished regions with different permeability properties. Additionally, the comparisons of the regional membrane oxygen permeability coefficients with that of water indicate which region forms a barrier or a pathway (“hydrophobic channels”) for oxygen transport This paper also establishes a methodological base for further investigations of oxygen transport into the human eye lens.