Regional differences in the lateral mobility of plasma membrane lipids in a molluscan embryo

Regional and temporal differences in plasma membrane lipid mobility have been analyzed during the first three cleavage cycles of the embryo of the polar-lobe-forming mollusc Nassarius reticulatus by the fluorescence photobleaching recovery (FPR) method, using 1,1'-ditetradecyl 3,3,3',3'-tetramethylindocarbocyanine iodide (C14diI) as a fluorescent lipid probe. During this period of development the lateral diffusion coefficient of membrane lipids is consistently greater in the vegetal polar lobe area as compared to the animal plasma membrane area (on average 30%), demonstrating the existence of an animal-vegetal polarity in plasma membrane properties. At third cleavage, the differences between animal and vegetal plasma membrane region become even more pronounced; in the four animal micromeres the diffusion coefficient (D) and mobile fraction (MF) are 2.9 +/- 0.2 X 10(-9) cm2/sec and 51 +/- 2%, respectively, while in the four vegetal macromeres D = 5.0 +/- 0.3 X 10(-9) cm2/sec and MF = 78 +/- 2%. Superimposed upon the observed animal-vegetal polarity, the lateral diffusion in the polar lobe membrane area shows a cell-cycle-dependent modulation. The highest mean values for D are reached during the S phase (ranging from 7.0 to 7.8 X 10(-9) cm2/sec in the three cycles measured), while at the end of G2 phase and during early mitosis mean values for D have decreased significantly (ranging from 5.0 to 5.9 X 10(-9) cm2/sec). Diffusion rates in the animal membranes of the embryo are constant during the three successive cell cycles (D = 4.3-5.0 X 10(-9) cm2/sec), except for a peak at the S phase of the first cell cycle (D = 6.0 X 10(-9) cm2/sec). These results are discussed in relation with previously observed ultrastructural heterogeneities in the Nassarius egg plasma membrane. It is speculated that the observed animal-vegetal polarity in the organization of the egg membrane might play an important role in the process of cell diversification during early development.     

Kinetics of the processes of desorption from fatty acid monolayers

The surface area, A, of contracting fatty acid monolayers was measured as a function of time, t, at constant surface pressure. In the initial temporal phase, ln A was linear with radical t. In a subsequent steady-state phase, ln A was linear with t. The initial desorption coefficient for sodium palmitate, K(i), and the steady-state desorption coefficient, K(s), varied directly with surface pressure and subphase pH, and these desorption coefficients also varied with the composition of the subphase buffer. However, the K(s)/K(i) ratio was independent of these variables. The diffusion coefficient, D(25), for sodium palmitate calculated from desorption coefficient ratios was 4.8 +/- 0.6 x 10(-6) cm(2)/sec. This value was in reasonable agreement with D(25) for sodium palmitate measured by time-lag diffusion, 3.7 +/- 0.6 x 10(-6) cm(2)/sec. D(25) values obtained for a series of fatty acids suggested that higher members of the series diffused as small aggregates averaging two to four molecules in size. Kinetic and diffusion data both supported a model for the desorption process described by Ter Minassian-Saraga.     

Lateral diffusion of lipids and glycophorin in solid phosphatidylcholine bilayers. The role of structural defects

The lateral mobility of the lipid analog N-4-nitrobenzo-2-oxa-1,3 diazole phosphatidylethanolamine and of the integral protein glycophorin in giant dimyristoylphosphatidylcholine vesicles was studied by the photobleaching technique. Above the temperature of the chain-melting transition (Tm = 23 degrees C), the diffusion coefficient, Dp, of the protein [Dp = (4 +/- 2) X 10(-8) cm2/s at 30 degrees C] was within the experimental errors equal to the corresponding values DL of the lipid analog. In the P beta 1 phase the diffusion of lipid and glycophorin was studied as a function of the probe and the protein concentration. (a) At low lipid-probe content (cL less than 5 mmol/mol of total lipid), approximately 20% of the probe diffuses fast (D approximately equal to 10(-8) - 10(-9) cm2/s), while the mobility of the rest is strongly reduced (D less than 10(-10) cm2/s). At a higher concentration (cp approximately 20 mmol), all probe is immobilized (D less than 10(-10) cm2/s). (b) Incorporation of glycophorin up to cp = 0.4 mmol/mol of total lipid leads to a gradual increase of the fraction of mobile lipid probe due to the lateral-phase separation into a pure P beta 1 phase and a fraction of lipid that is fluidized by strong hydrophilic lipid-protein interaction. (c) The diffusion of the glycophorin molecules is characterized by a slow and a fast fraction. The latter increases with increasing protein content, which is again due to the lateral-phase separation caused by the hydrophilic lipid-protein interaction. The results are interpreted in terms of a fast transport along linear defects in the P beta 1 phase, which form quasi-fluid paths for a nearly one dimensional and thus very effective transport. Evidence for this interpretation of the diffusion measurements is provided by freeze-fracture electron microscopy.     

Two-dimensional electron transfer from cytochrome C to photosynthetic reaction centers

The arrangement and the electron transfer are studied for photosynthetic reaction centers (RC) of Rhodopseudomonas sphaeroides reconstituted into phospholipid vesicles. Freeze-etch electron micrographs of phase separated mixed vesicles reveal an RC enrichment in the phase containing the acidic lipid serine. It is demonstrated that the electron transfer from cytochrome c to RC involves a two-dimensional diffusion of the membrane bound electron donor with diffusion coefficients (D approximately 10(-9) cm2/sec) characteristic for membrane proteins.     

Rapid lateral diffusion of the variant surface glycoprotein in the coat of Trypanosoma brucei

The membrane form of the variant surface glycoprotein (mfVSG) is anchored in the plasma membrane of Trypanosoma brucei by a dimyristoylphosphatidylinositol residue connected via a glycan to the COOH-terminal amino acid. The glycoprotein molecules are tightly packed, forming a coat that is impenetrable to lytic serum components. Lateral diffusion of mfVSG was measured by the fluorescence recovery after photobleaching technique. mfVSG labeled on the cell surface with rhodamine-conjugated anti-VSG Fab fragments showed a diffusion coefficient of 1 X 10(-10) cm2/s at 37 degrees C and of 0.7 X 10(-10) cm2/s at 27 degrees C. About 80% of the molecules were mobile. Affinity-purified mfVSG molecules implanted into the plasma membrane of baby hamster kidney cells exhibited a similar mobility to that found in the trypanosome coat [D = (0.4-0.7) X 10(-10) cm2/s at 4 degrees C]. Phospholipid mobility in the plasma membrane of trypanosomes was characterized by a diffusion coefficient of 2.2 X 10(-9) cm2/s at 37 degrees C. It is concluded that mfVSG mobility in the surface coat of the parasite is rapid and comparable to that of other membrane-bound glycoproteins but slower than that of phospholipids.     

Non-steady-state diffusion in a multilayered tissue initiated by manipulation of chemical activity at the boundaries.

Diffusion of ionic and nonionic species in multilayered tissues plays an important role in the metabolic processes that take place in these tissues. To create a mathematical model of these diffusion processes, we have chosen as an example hydrogen-bicarbonate ion pair diffusion within the mammalian cornea. This choice was based on the availability of experimental data on this system. The diffusion coefficient of the hydrogen-bicarbonate ion pair in corneal stroma and epithelium is calculated from the observed change in pH in the stroma when conditions at the corneal anterior epithelial surface are changed while the posterior surface is continually bathed with a Ringer's solution in equilibrium with a CO2-gas air mixture. Matching experimental results to a mathematical model of the cornea as a two-layer diffusion system yields, at 37 degrees C, a diffusion coefficient of the hydrogen-bicarbonate ion pair of 2.5 x 10(-6) cm2/s in the stroma and 0.4 x 10(-6) cm2/s in the epithelium. Application of the Nernst-Einstein equation to these data gives the following diffusion coefficients in the two layers: 1) stroma, D(H+) = 11.8 x 10(-6) cm2/s; D(HCO3-) = 1.5 x 10(-6) cm2/s; and 2) epithelium, D(H+) = 1.9 x 10(-6) cm2/s; D(HCO3-) = 0.22 x 10(-6) cm2/s.     

The purification and properties of isocitrate lyase from Chlorella

1. Isocitrate lyase (threo-d(s)-isocitrate glyoxylate-lyase, EC 4.1.3.1) has been purified from acetate-adapted cells of Chlorella pyrenoidosa. 2. The final preparation was homogeneous by the criteria of sedimentation, diffusion and polyacrylamide-gel electrophoresis. 3. The sedimentation coefficient (S(20,w)) was 9.04x10(-13)sec. and the diffusion coefficient (D(20,w)) 4.62x10(-7)cm.(2)/sec.; from these values the molecular weight of the enzyme was calculated to be 170000 and its Stokes radius to be 4.63x10(-7)cm. 4. The elution of the enzyme from Sephadex G-100 was studied and estimates of molecular weight and Stokes radius were obtained from the elution data. 5. The turnover number of the enzyme was 5950moles of glyoxylate formed/min./mole of enzyme at 30 degrees . 6. With threo-d(s)(+)-isocitrate as substrate, the K(m) of the enzyme was 0.023mm.     

Lateral mobility of plasma membrane lipids in dividing Xenopus eggs

The lateral mobility of plasma membrane lipids was analyzed during first cleavage of Xenopus laevis eggs by fluorescence photobleaching recovery (FPR) measurements, using the lipid analogs 5-(N-hexadecanoyl)aminofluorescein ("HEDAF") and 5-(N-tetradecanoyl)aminofluorescein ("TEDAF") as probes. The preexisting plasma membrane of the animal side showed an inhomogeneous, dotted fluorescence pattern after labeling and the lateral mobility of both probes used was below the detection limits of the FPR method (D much less than 10(-10) cm2/sec). In contrast, the preexisting plasma membrane of the vegetal side exhibited homogeneous fluorescence and the lateral diffusion coefficient of both probes used was relatively high (HEDAF, D = 2.8 X 10(-8) cm2/sec; TEDAF, D = 2.4 X 10(-8) cm2/sec). In the cleaving egg visible transfer of HEDAF or TEDAF from prelabeled plasma membrane to the new membrane in the furrow did not occur, even on the vegetal side. Upon labeling during cleavage, however, the new membrane was uniformly labeled and both probes were mobile, as in the vegetal preexisting plasma membrane. These data show that the membrane of the dividing Xenopus egg comprises three macrodomains: (i) the animal preexisting plasma membrane; (ii) the vegetal preexisting plasma membrane; (iii) the new furrow membrane.     

Alpha interferon accelerates lateral diffusion of Daudi cell surface differentiation antigens: measurement by fluorescence redistribution after photobleaching

Lateral diffusion coefficients (D) of two surface differentiation antigens (sIgM and Bp35) were determined on interferon-sensitive (-IFs) or resistant (-IFr) Daudi cells by fluorescence photobleaching, using monospecific FITC-anti-IgM or PE-anti-Leu 16 probes. For untreated Daudi -IFs, mean (D) were 5.8 and 5.3 (x10(-10) cm2/sec). These increased, to 11 and 7.9 x 10(-10) cm2/sec (p less than 0.001) within 30 min after binding of recombinant IFN-a (80 to 800 U/10(6) cells), but decreased by up to 4-fold after Con A Mean (D) of identical surface antigens on Daudi-IFr were 8.2 and 9.4 x 10(-10) cm2/sec; and were not altered by IFN-a. Mean (D) of a lipid analog was up to 40-fold higher than for surface proteins and statistically identical in Daudi-IFs and Daudi-IFr. Rapid acceleration by IFN-a of surface protein lateral diffusion in Daudi-IFs obviously could facilitate anti-proliferative signal transduction; by contrast, a baseline increase of (D) in Daudi-IFr was evidently associated with their refractory state.     

Oxygen diffusion in biological and artificial membranes determined by the fluorochrome pyrene

Quenching of pyrene fluorescence by oxygen was used to determine oxygen diffusion coefficients in phospholipid dispersions and erythrocyte plasma membranes. The fluorescence intensity and lifetime of pyrene in both artificial and natural membranes decreases about 80% in the presence of 1 atm O2, while the fluorescence excitation and emission spectra and the absorption spectrum are unaltered. Assuming the oxygen partition coefficient between membrane and aqueous phase to be 4.4, the diffusion coefficients for oxygen at 37 degrees C are 1.51 X 10(-5) cm2/s in dimyristoyl lecithin vesicles, 9.32 X 10(-6) cm2/s in dipalmitoyl lecithin vesicles, and 7.27 X 10(-6) cm2/s in erythrocyte plasma membranes. The heats of activation for oxygen diffusion are low (less than 3 kcal/degree-mol). A dramatic increase in the diffusion constant occurs at the phase transition of dimyristoyl and dipalmitoyl lecithin, which may result from an increase in either the oxygen diffusion coefficient, partition coefficient, or both. The significance of the change in oxygen diffusion below and above the phase transition for biological membranes is discussed.     

Relationship between water transfers and the salinity of the environment in the isopod crustacean Sphaeroma serratum (Fabricius)]

The isopod Crustacean Sphaeroma serratum is isotonic to the medium in sea water and hypertonic in diluted media. The drinking rate is 15.9 microliter in SW and 3.2 microliter 24 h-1 100 mg-1 wet weight in 50% SW. The extracellular space is 28.4% in SW and 27.0% of the wet weight in 50% SW. The rate of urine production, calculated from the half time for the loss of sodium diatriazoate is 11.8 mg in SW and 42.9 mg 24 h-1 100 mg-1 wet weight in 50% SW. 95% of the diffusion fluxes of water take place through the pleopods: their surface is about 62.6 mm2 in a 100 mg weighing animal. The activation energy of water molecules is 15.0 kcal/mol between +5 degrees C and +15 degrees C and 6.7 kcal/mol between +15 degrees C and +25 degrees C. The diffusion permeability coefficient Pd is 1.71 X 10(-4) cm/sec in SW and 0.70 X 10(-4) cm/sec in 50% SW. The osmotic permeability coefficient Pos has a mean value of 1.91 X 10(-4) cm/sec in SW and 1.24 X 10(-4) cm/sec in 50% SW. Our results are compared with those obtained in other Crustaceans and in Fishes. Their validity is discussed (influence of unstirred layers, solute-solvent interaction). They are explained according to the different theories dealing with the water transit through the membranes.     

Bacteriophage phiNS11: a lipid-containing phage of acidophilic thermophilic bacteria. IV. Sedimentation coefficient, diffusion coefficient, partial specific volume, and particle weight of the phage.

The particle weight (molecular weight) of phiNS11 was determined from the sedimentation coefficient, diffusion coefficient, and partial specific volume of the phage. The sedimentation coefficient of the phage (S(0)20, W) is 416 +/- 2.7S. The diffusion coefficient D(0)20, W), which was determined by quasielastic light scattering measurement, is (0.57 +/- 0.03) x 10(-7) cm2/s. The partial specific volume was determined by the mechanical oscillation technique to be 0.747 +/- 0.007 cm3/g. Based on these values, the particle weight of the phage was calculated to be (70.3 +/- 4.3) x 10(6) daltons, which agrees well with the particle weight (69--72 x 10(6) daltons) estimated from the molecular weight of phage DNA and the content of DNA. The Stokes radius of the phage particle was calculated to be 37.7 +/- 2 nm and hydration of the phage was estimated to be 1.18 cm3/g of dry phage. From the particle weight and the chemical composition of the phage, we estimated that one phage particle contains one double-stranded DNA molecule, 16,000 residues of fatty acid, 72 protein I molecules, 920 protein II, 42 protein III, 48 protein IV, 290 protein V molecules, and 3,700 molecules of polyamines.     

Characterization of interaction between cationic lipid-oligonucleotide complexes and cellular membrane lipids using confocal imaging and fluorescence correlation spectroscopy

Complexes formed by cationic liposomes and single-strand oligodeoxynucleotides (CL-ODN) are promising delivery systems for antisense therapy. ODN release from the complexes is an essential step for inhibiting activity of antisense drugs. We applied fluorescence correlation spectroscopy and confocal laser scanning microscopy to monitor CL-ODN complex interaction with membrane lipids leading to ODN release. To model cellular membranes we used giant unilamellar vesicles and investigated the transport of Cy-5-labeled ODNs across DiO-labeled membranes. For the first time, we directly observed that ODN molecules are transferred across the lipid bilayers and are kept inside the giant unilamellar vesicles after release from the carriers. ODN dissociation from the carrier was assessed by comparing diffusion constants of CL-ODN complexes and ODNs before complexation and after release. Freely diffusing Cy-5-labeled ODN (16-nt) has diffusion constant D(ODN) = 1.3 +/- 0.1 x 10(-6) cm2/s. Fluorescence correlation spectroscopy curves for CL-ODN complexes were fitted with two components, which both have significantly slower diffusion in the range of D(CL-ODN) = approximately 1.5 x 10(-8) cm2/s. Released ODN has the mean diffusion constant D = 1.1 +/- 0.2 x 10(-6) cm2/s, which signifies that ODN is dissociated from cationic lipids. In contrast to earlier studies, we report that phosphatidylethanolamine can trigger ODN release from the carrier in the full absence of anionic phosphatidylserine in the target membrane and that phosphatidylethanolamine-mediated release is as extensive as in the case of phosphatidylserine. The presented methodology provides an effective tool for probing a delivery potential of newly created lipid formulations of CL-ODN complexes for optimal design of carriers.     

Effect of fatty acids on the membrane fluidity of cultured chick dorsal root ganglion measured by fluorescence photobleaching recovery

The effect of fatty acids on the membrane fluidity in tissue cultured chick embryo dorsal root ganglion was studied by fluorescence recovery method. Lateral motion of the lipid was measured by observing the fluorescent probe, 5-(octadecylthiocarbamoylamino) fluorescence, F18. The effective lateral diffusion coefficient of the membrane was around 0.30 X 10(-8) cm2/sec in control cells, 0.42 X 10(-8) cm2/sec in 2-decenoic acid treated cells, and 0.35 X 10(-8) cm2/sec in valeric acid treated cells. From these results it is concluded that effective mobilities of the membrane complex increased about 40% by the external application of 2-decenoic acid, while valeric acid increased it only 12%. From the physiological results that 2-decenoic acid inhibits the Na-channel, it is suggested that this increase in the membrane fluidity might affect the Na-channel.     

Rapid diffusion of spectrin bound to a lipid surface.

Human erythrocyte spectrin was labelled with the probe 5, 5'-disulfato-1-(6-hexanoic acid N-hydroxysuccinimide ester)-1'-ethyl-3,3,3',3'-tetramethylindocarbocyanine (Cy3). Cy3-spectrin was bound to the outer surface of dimyristoylphosphatidylcholine (DMPC) multilamellar vesicles and its diffusion measured by fluorescence recovery after photobleaching (FRAP). It was found that at 30 degrees C, above the lipid gel to liquid-crystalline phase transition of the lipids, Cy3-spectrin had an unexpectedly high diffusion coefficient D=(2.1+/-0.6)x10(-7)) cm2/s. At the phase transition, diffusion of Cy3-spectrin was only slightly lower; D=(1.3+/-0.3)x10(-7) cm2/s, whereas at 14 degrees C, well below the lipid phase transition, diffusion was found to be much slower with D=(3.1+/-0.12)x10(-9) cm2/s. The fast diffusion of Cy3-spectrin on the lipid surface implies that the individual bonds which bind spectrin to the lipid surface must rapidly be made and broken. In the light of these results, spectrin-lipid interactions alone appear unlikely to have any significant role in supporting the cell membrane. Probably, the interactions serve only to localise the spectrin at the inner lipid surface in order to facilitate formation of the cytoskeleton.     

Growth mechanism of myelin figures of phosphatidylcholine

The initial growth process of myelin figures, rod-like lyotropic liquid-crystalline structures, formed by phosphatidylcholine in water, ethylene glycol or glycerin, is suggested to be diffusion-limited with an apparent diffusion coefficient D of approx. 10(-6) cm2/s. D can be expressed by the sum of two processes. One is considered to describe the diffusion of an aggregate of phosphatidylcholine molecules and the other mainly to describe a lateral diffusion in the bilayer membranes which constitute myelin figures.     

Surface properties of membrane vesicles prepared from muscle cells of Ascaris suum.

To facilitate biochemical, pharmacological, and biophysical studies on the membrane of the body muscle of Ascaris suum, a method for preparing intact vesicles was developed. Vesicles were prepared by incubating a muscle flap preparation with 1 mg/ml collagenase in a saline solution and then washing in saline without enzyme. The vesicles then formed gradually over the next hour as outgrowths of the original surface membrane from the bag region of the muscle. The vesicles were harvested readily by suction using a Pasteur pipette. The structure of the vesicles was examined with the transmission electron microscope. The whole-cell patch-clamp technique showed that the vesicles had a high input resistance and that the membrane was complete. The vesicle membrane was shown to contain Ca-activated Cl channels and gamma-aminobutyric acid-activated Cl channels. The vesicles also were shown to be suitable for fluorescence recovery after photobleaching studies designed to examine lateral and vertical movement of a lipid probe (5-N [octadecanoyl]-aminofluorescein) in the membrane. This probe had a mean lateral diffusion coefficient (DL) of 8.1 x 10(-9) cm2/sec, but only a proportion (68.4%) of the probe was mobile. The latter observation illustrated the nonuniform nature of the membrane. Ivermectin (10(-7) M) had no effect on DL or percent recovery. Trypan blue quenching experiments showed that the lipid probe remained in the outer monolayer of the membrane. These observations illustrate the experimental value of the vesicles; they are potentially useful in discerning anthelmintic mode of action and in drug screening.     

Binding of ethidium to DNA measured using a 2D diffusion-modulated gradient COSY NMR experiment

The binding of ethidium bromide to a DNA hairpin (dU(5)-hairpin) was investigated using a novel 2D diffusion-modulated gradient correlation spectroscopy (DMG-COSY) experiment to evaluate the applicability of this technique for studying the binding of drugs to DNA. The DMG-COSY experiment includes a preparation period during which coherent magnetization is attenuated due to molecular self-diffusion. Magnetization then evolves due to scalar coupling during an evolution delay, and is detected using gradient pulses for coherence selection. The time-domain data are processed in an analogous manner as for gradient-selected COSY experiments. The diffusion coefficient for uridine in DMSO solution was determined from the H5-H6 crosspeak intensities for a series of 2D DMG-COSY experiments that differed in the magnitude of the gradient pulses applied during the preparation period of the DMG-COSY experiment. The diffusion coefficient for uridine calculated from the DMG-COSY experiments was identical (within experimental error) to that determined from 1D diffusion experiments (5.24x10(-6) cm(2)/s at 26 degrees C). The diffusion coefficients for ethidium bromide and for the dU(5)-hairpin were first measured separately using the DMG-COSY experiment, and then measured in the putative complex. The diffusion coefficient for free ethidium bromide (4.15x10(-6) cm(2)/s at 26 degrees C) was considerably larger than for the dU(5)-hairpin (1. 60x10(-6) cm(2)/s at 26 degrees C), as expected for the smaller molecule. The diffusion coefficient for ethidium was markedly decreased upon addition of the dU(5)-hairpin, consistent with complex formation (1.22x10(-6) cm(2)/s at 26 degrees C). Complex formation of 1:1 stoichiometry between ethidium and the stem of the dU(5)-hairpin was verified independently by fluorescence spectroscopy. These results demonstrate the utility of the DMG-COSY experiment for investigating the binding of drugs to DNA in aqueous solution.     

Surface diffusion in human serum lipoproteins

From the viscosity dependence of the 31P NMR signals, the diffusion coefficients DT of phospholipid molecules in the surface monolayer of HDL, LDL and VLDL have been determined. DT for HDL3 and HDL2 are found to be 2.3 X 10(-8) cm2/s and 1.8 X 10(-8) cm2/s, respectively. These values are similar to values reported for diffusion of phospholipid molecules in phospholipid bilayers above the gel to liquid crystalline phase transition temperature. Viscosity dependence of [16,16,16-2H3]phosphatidylcholine incorporated into HDL2 yielded a value similar to that determined by 31P (DT = 1.9 X 10(-8) cm2/s). Slower diffusion coefficients were measured for LDL2 and VLDL. VLDL had a value DT = 9.1 X 10(-9) cm2/s. The diffusion coefficient for LDL2 was 1.4 X 10(-9) cm2/s. Thus, diffusion of phospholipids in LDL2 is a full order of magnitude slower at 25 degrees C than diffusion of phospholipids in the HDLs.     

Diffusion and consumption of oxygen in the superfused retina of the drone (Apis mellifera) in darkness

Double-barreled O2 microelectrodes were used to study O2 diffusion and consumption in the superfused drone (Apis mellifera) retina in darkness at 22 degrees C. Po2 was measured at different sites in the bath and retinas. It was found that diffusion was essentially in one dimension and that the rate of O2 consumption (Q) was practically constant (on the macroscale) down to Po2 s less than 20 mm Hg, a situation that greatly simplified the analysis. The value obtained for Q was 18 +/- 0.7 (SEM) microliter O2/cm3 tissue . min (n = 10), and Krogh's permeation coefficient (alpha D) was 3.24 +/- 0.18 (SEM) X 10(-5) ml O1/min . atm . cm (n = 10). Calculations indicate that only a small fraction of this Q in darkness is necessary for the energy requirements of the sodium pump. the diffusion coefficient (D) in the retina was measured by abruptly cutting off diffusion from the bath and analyzing the time-course of the fall in Po2 at the surface of the tissue. The mean value of D was 1.03 +/- 0.08 (SEM) X 10(-5) cm2/s (n = 10). From alpha D and D, the solubility coefficient alpha was calculated to be 54 +/- 4.0 (SEM) microliter O2 STP/cm3 . atm (n = 10), approximately 1.8 times that for water.