The osmotic behavior of corn mitochondria

The volume changes undergone by corn (Zea mays L.) mitochondria suspended in solutions of constant or varying osmolarity were studied. Within the range of osmotic pressure from 1.8 to 8.4 atmospheres, corn mitochondria behave as osmometers, if allowance is made for an osmotic “dead space” of about 6.9 μl/mg protein. The final equilibrium volume of mitochondria swollen in solutions containing both ribose and sucrose were shown to depend upon the concentration of impermeable solute (sucrose) present and not upon the concentration of ribose present. Osmotic reversibility was found for mitochondria swollen in isotonic solutions of KCI or ribose. The passive swelling of corn mitochondria may be due to the osmotic flow of water coupled to the diffusion of a permeable solute.     

Mitochondria in the flight muscles of insects. III. Mitochondrial cytochrome c in relation to the aging and wing beat frequency of flies

1. In the present study a correlation has been sought between aging, flight muscle mitochrondria (sarcosomes), cytochrome c, and flight ability in the blowfly, Phormia regina. 2. During the 1st week of adult life, individual sarcosomes increase in mass from 2.7 x 10^–7 µg. dry weight at the time of emergence, to 8.5 x 10^–7 µg. by the 7th day. During this period of growth, the number of sarcosomes per fly (6.7 x 10⁸) remains constant. When mature, the sarcosomes account for 32.6 per cent of the total muscle dry weight, or close to 40 per cent on a wet weight basis. 3. It appears probable that the high content of flight muscle cytochromes is entirely localized in the sarcosomes. The cytochromes continue to be synthesized and increase in titer within the sarcosomes for 7 days after adult emergence. 4. As determined spectroscopically, the various cytochrome components at all times maintain a constant ratio both to one another and to the sarcosomal dry weight. This suggests the possibility that the cytochrome system may be synthesized as a single entity. 5. The wing-beat frequency of Drosophila funebris and Phormia varies with the age of these flies, being lowest at the time of emergence and maximum after the 6th day. 6. The relations between wing-beat frequency, respiration during flight, and sarcosomal cytochrome c content are discussed. On the basis of some likely assumptions it is calculated that the cytrochrome c turnover number is over 5,000, and that the cytochrome c turns over once for every two wing-beat cycles.     

Cytochrome P-450 and hydroxylating activity of microsomal preparations from whole houseflies

1. A new microsomal preparation, obtained from whole houseflies is described in terms of its cytochrome P-450 content and its hydroxylating activity. 2. Microsomes prepared from whole-fly brei, obtained with the aid of a mortar (a procedure that avoids the destruction of sarcosomes), contain 0.265nmol of cytochrome P-450 and hydroxylate naphthalene at a rate of 28.5nmol/mg of microsomal protein in 30min at 30 degrees C. This corresponds to 104nmol of naphthalene hydroxylated/nmol of cytochrome P-450. This is the highest rate ever reported for housefly and rat liver microsomal preparations. 3. Microsomal fractions prepared by procedures that do not retain the integrity of sarcosomes show the presence in the CO-difference spectrum of a 428nm peak. This cytochrome is associated with sarcosomal microsomes and it may be involved in the inhibition of insect microsomal mixed-function oxidases, although other factors cannot be discarded at present. 4. The inability to show cytochrome P-450 in microsomal fractions isolated from whole houseflies by other procedures may be at least partially due to a masking effect brought about by contamination with the sarcosomal cytochrome.     

Ultrastructural changes in sarcosomes after exposure of adult Calliphora erythrocephala to lethal and sub-lethal high temperatures

The effects of lethal and sub-lethal high temperatures on the morphology of intact flight muscle sarcosomes of adult Calliphora erythrocephala are described. Treatment of adult flies to lethal temperatures results in ultrastructural changes in the organisation of the cristae and the appearance of electron opaque inclusions. These changes have not been observed after sub-lethal heat treatment, when 50% of the animals recover. It is suggested that changes in the ultrastructure of the intact sarcosomes may be correlated with changes in their ability to couple oxidative phosphorylation with α-glycerophosphate. Age-dependent changes in the sensitivity of sarcosomes are related to changes in the heat death point of the animal and suggest that the impairment of sarcosomal function may be one of the primary lesions in heat death of adult C. erythrocephala.     

The mechanism of mitochondrial swelling. VIII. Permeability of mitochondria to alkali metal acetates

The capacity of beef heart mitochondria to undergo osmotically induced volume changes in decimolar M⁺-acetate or other weak acid anion media is characterized by the following features: (1) mitochondria resist swelling when suspended in potassium or rubidium acetate media in the presence of respiratory inhibitors; (2) mitochondria swell extensively when suspended in ammonium or sodium acetate media in the presence of respiratory inhibitors; and (3) actively respiring mitochondria swell extensively whether suspended in ammonium, sodium, potassium, or rubidium acetate media. These findings have been interpreted to mean that (1) the nonenergized mitochondrial inner membrane is permeable to acetate anions, (2) the nonenergized mitochondrial inner membrane is permeable to ammonium and sodium ions in the presence of acetate or other weak acid anions, (3) the nonenergized mitochondrial inner membrane is relatively impermeable to potassium and rubidium ions in the presence of acetate or other weak acid anions, and (4) energized mitochondria are considerably more permeable to potassium and rubidium (acetate) ions than are non-energized mitochondria. The experiments described in this communication which provide the evidence for these interpretations involve methods which are independent of volume changes. The results confirm the first three of the above interpretations but are inconsistent with the fourth. A general theory for passive ion movements in mitochondria is presented and the results are discussed in terms of the development of an energy dependent ion gradient as the key to energized swelling in potassium or rubidium acetate.     

THE STRUCTURE OF THE COLLODION MEMBRANE AND ITS ELECTRICAL BEHAVIOR : IX. WATER UPTAKE AND SWELLING OF COLLODION MEMBRANES IN AQUEOUS SOLUTIONS OF ORGANIC ELECTROLYTES AND NON-ELECTROLYTES

1. Dried collodion membranes are known to swell in water and to the same limited extent also in solutions of strong inorganic electrolytes (Carr and Sollner). The present investigation shows that in solutions of organic electrolytes and non-electrolytes, the swelling of dried collodion membranes is not as uniform, but depends on the nature of the solute. 2. The solutions of typically "hydrophilic" substances, e.g., glycerine, glucose, and citric acid, swell collodion membranes only to the same extent as water and solutions of strong electrolytes. In solutions of typically carbophilic substances (e.g., butyric acid, valeric acid, isobutyl alcohol, valeramide, phenol, and m-nitrophenol) the swelling of the membranes is much stronger than in water, according to the concentration used. For the brand of collodion used the swelling in 0.5 M solution was in some cases as high as 26 per cent of the original volume, as compared to 6 to 7 per cent in water. Therefore, in these solutions the "water-wetted dried" collodion membrane is not rigid, inert, and non-swelling, but behaves as a swelling membrane. 3. The solutes which cause an increased swelling of the membranes are accumulated in the latter, the degree of accumulation being markedly parallel with the degree of their specific swelling action. 4. The anomalously high permeabilities of certain carbophilic organic solutes reported by Michaelis, Collander, and Höber find an explanation in the specific interaction of these substances with collodion. 5. The use of the collodion membrane as a model of the ideal porous membrane is restricted to those instances in which no specific interaction occurs between the solute and the collodion.     

Osmotic sensitivity of canine spermatozoa

The objective of this study was to determine osmotic tolerance of canine spermatozoa. The study comprised three experiments: (1) spermatozoa suspended either in an egg yolk-citrate (EYC) extender or in Kenney skim milk extender were exposed to NaCl solutions ranging from 290 to 1500 mOsm; (2) spermatozoa suspended in EYC were exposed to 550 to 1500 mOsm solutions of glucose, galactose, or fructose; and (3) spermatozoa suspended in EYC or glucose-bovine serum albumin (G-BSA) were exposed to 0.6 M (approximately 900 mOsm) or 1.2 M (approximately 1600 mOsm) solutions of glycerol, ethylene glycol (EG), or dimethyl sulfoxide (Me(2)SO). In all experiments, motility and membrane integrity of spermatozoa were assessed after they were diluted into isotonic medium at 37 degrees C. Exposure of canine spermatozoa to solutions of either NaCl or monosaccharides at osmolalities >500 mOsm caused a significant reduction of motility (P<0.01). Motility of spermatozoa was more affected by osmotic stress than their membrane integrity. Osmotic sensitivity of canine spermatozoa was dependent on the type of extender; spermatozoa suspended in the Kenney extender were more resistant to osmotic stress than those suspended in the EYC extender. Despite their sensitivity to exposure to high concentrations of nonpermeating agents, canine spermatozoa were rather resistant to exposure to glycerol and EG. However, Me(2)SO was toxic to canine spermatozoa; motility was substantially reduced after spermatozoa were exposed to 0.6 M Me(2)SO. The type of extender also affected the sensitivity of canine spermatozoa to Me(2)SO; spermatozoa suspended in the EYC extender were more resistant than those suspended in G-BSA. It was concluded that canine spermatozoa are sensitive to osmotic stress, but are tolerant to shrinking and swelling caused by exposure to permeating cryoprotectants.     

THE RECIPROCAL RELATION BETWEEN THE OSMOTIC PRESSURE AND THE VISCOSITY OF GELATIN SOLUTIONS

1. These experiments confirm the conclusion that protein solutions are true solutions consisting of isolated ions and molecules, and that these solutions may or may not contain in addition solid submicroscopic particles capable of occluding water. 2. The typical influence of electrolytes on the osmotic pressure of protein solutions is due to the isolated protein ions since these alone are capable of causing a Donnan equilibrium across a membrane impermeable to the protein ions but permeable to most crystalloidal ions. 3. The similar influence of electrolytes on the viscosity of protein solutions is due to the submicroscopic solid protein particles capable of occluding water since the amount of water occluded by (or the amount of swelling of) these particles is regulated by the Donnan equilibrium. 4. These ideas are supported by the fact that the more the submicroscopic solid particles contained in a protein solution or suspension are transformed into isolated ions (e.g., by keeping gelatin solution for 1 hour or more at 45°C.) the more the viscosity of the solution is diminished while the osmotic pressure is increased, and vice versa.     

Fusion of phospholipid vesicles with a planar membrane depends on the membrane permeability of the solute used to create the osmotic pressure

Phospholipid vesicles fuse with a planar membrane when they are osmotically swollen. Channels in the vesicle membrane are required for swelling to occur when the vesicle-containing compartment is made hyperosmotic by adding a solute (termed an osmoticant). We have studied fusion using two different channels, porin, a highly permeable channel, and nystatin, a much less permeable channel. We report that an osmoticant's ability to support fusion (defined as the magnitude of osmotic gradient necessary to obtain sustained fusion) depends on both its permeability through lipid bilayer as well as its permeability through the channel by which it enters the vesicle interior. With porin as the channel, formamide requires an osmotic gradient about ten times that required with urea, which is approximately 1/40th as permeant as formamide through bare lipid membrane. When nystatin is the channel, however, fusion rates sustained by osmotic gradients of formamide are within a factor of two of those obtained with urea. Vesicles containing a porin-impermeant solute can be induced to swell and fuse with a planar membrane when the impermeant bathing the vesicles is replaced by an isosmotic quantity of a porin-permeant solute. With this method of swelling, formamide is as effective as urea in obtaining fusion. In addition, we report that binding of vesicles to the planar membrane does not make the contact region more permeable to the osmoticant than is bare lipid bilayer. In the companion paper, we quantitatively account for the observation that the ability of a solute to promote fusion depends on its permeability properties and the method of swelling. We show that the intravesicular pressure developed drives fusion.     

Hypothermic preservation of hepatocytes : I. Role of cell swelling

Hepatocytes from isolated rat livers were hypothermically incubated (5 degrees C) in an oxygenated environment with continuous shaking (to simulate organ perfusion preservation). The incubation solution was either a tissue culture medium (L-15), an organ preservation perfusate (UW gluconate), or a simple cold-storage solution used for organ preservation (UW lactobionate). Hepatocyte viability was assessed from the release of lactate dehydrogenase (LDH) into the incubation medium. Cell swelling (due to the uptake of water) was also measured. Within 24 hr, hepatocytes hypothermically stored in each of the three incubation solutions became swollen (30 to 40% water gain) and lost a significant amount of LDH (as much as 60%). The addition of polyethylene glycol (PEG; relative molecular mass 8000; 5 g%) to the solutions suppressed cell swelling and allowed the incubated hepatocytes to remain relatively well preserved (30% LDH release) for as long as 120 hr. Adding either dextran (relative molecular mass 10,000 to 78,000; 5 g%) or saccharides (100 mmol/liter) instead of PEG neither prevented cell swelling nor prevented the cells from dying. The results of this study suggest (i) there is a direct correlation (r = 0.873) between hypothermia-induced cell swelling and cell death (i.e., the suppression of cell swelling prevents cell death); (ii) the mechanism by which PEG prevents cell swelling (and thus maintains cell viability) is not related to the osmotic or oncotic properties of the molecule but instead is apparently related to some unknown interaction between PEG and the cell, an interaction that provides stability during hypothermic incubation; and (iii) hypothermia-induced cell swelling must be prevented if isolated hepatocytes are to be used as a model for studying the mechanism by which cell damage occurs during hypothermic organ preservation. By eliminating cell death due to cell swelling, the biochemical mechanisms of cell death can be studied.     

Hemolysis of human red blood cells by freezing and thawing in solutions containing polyvinylpyrrolidone: relationship with postthypertonic hemolysis and solute movements

An investigation was made into the effects of the presence of polyvinylpyrrolidone (PVP) on changes in human red blood cells suspended in hypertonic solutions, on posthypertonic hemolysis, and on freezing at temperatures down to ?12 °C.PVP is very effective at reducing hemolysis when the red blood cells are frozen at temperatures down to ?12 °C. However, the membranes of the cells recovered on thawing have become very permeable to sodium and potassium ions and there is a much increased hemolysis if the cells are resuspended in an isotonic solution of sodium chloride.The presence of PVP does not affect the dehydration of the cells or the development of a change in membrane permeability when the cells are shrunken in hypertonic solutions at 0 °C. Neither does its presence in the hypertonic solution reduce the extent of posthypertonic hemolysis at 4 °C (as measured by the hemolysis on resuspension in an isotonic solution of sodium chloride), but it is more effective than sucrose at reducing hemolysis when present in the resuspension solution. It is concluded that the PVP is able to prevent swelling and hemolysis of cells which are very permeable to cations by opposing the colloid osmotic pressure due to the hemoglobin. However, this does not explain how PVP is able to protect cells against freezing damage at high cooling rates, and a mechanism by which it might do this is discussed.     

Coagulation of sea urchin egg cytoplasm by high salt concentrations

Unfertilized sea urchin eggs exposed to hyperosmotic salt solutions in excess of 1.75 M undergo a form of intracellular coagulation known as black cytolysis, similar to that seen in eggs injured by freezing. The process can be simulated by the microinjection of hypertonic salt into the cell suspended in isotonic solution in the absence of volume reduction. Black cytolysis during hyperosmotic stress can be attributed to the entry of concentrated extracellular solution through a membrane made permeable by excessive osmotic stress.     

Studies on the biochemical basis of susceptibility and resistance of the housefly to fluoroacetate

1. Administration of fluoroacetate to sensitive houseflies in amounts close to the L.D.₅₀ range (0.25–0.3 μg/fly) brought about a prompt elevation of their citrate content. With about 10-fold higher doses of fluoroacetate a concurrent increase of both citrate and pyruvate levels took place in the fly tissues.

2. Incubation of sarcosomes of the sensitive housefly strain in the presence of oxidizable substrates and fluoroacetate resulted in accumulation of citrate, inhibition of respiration and uncoupling of oxidative phosphorylation. The magnitude of the effects varied considerably with the different substrates used, being particularly pronounced with pyruvate and malate and inappreciable with succinate and -glycerophosphate.

3. The respiratory inhibition induced by a brief exposure in the cold of housefly sarcosomes to fluoroacetate, persisted after the sarcosomes had been washed free from fluoroacetate. The toxic effect of fluoroacetate on the respiratory chain could be prevented by an excess of simultaneously added acetate.

4. The susceptibility of the respiratory function of the sarcosomes to fluoroacetate inhibition was abolished by sonication. The unresponsiveness of the sonicated sarcosomes to fluoroacetate was attended by a loss of their respiratory chain phosphorylation activity.

5. Sarcosomes derived from a partially resistant housefly strain, when incubated in the presence of fluoroacetate, failed to accumulate citrate, but displayed the characteristic respiratory-inhibition response. Sarcosomes from a highly resistant strain showed no impairment of their functional capacity by fluoroacetate. However, all the different housefly strains tested proved to be equally sensitive to the deleterious effect of fluorocitrate on sarcosomal respiration.

6. The possible biochemical mechanisms underlying the toxicity of fluoroacetate in the housefly are considered with particular reference to the altered response of the target systems exhibited by the fluoroacetate-resistant strains.     

Thermal denaturation of the erythrocyte cytoskeleton alters the morphological changes associated with osmotic swelling

1. 1. The shape changes during osmotic swelling of human erythrocytes in a hypotonic medium at room temperature, at 45°C and at the denaturation temperature (49.5°C) of the cytoskeletal protein, spectrin, have been monitored by video microscopy.

2. 2. At room temperature the great majority of cells (which were discoid prior to injection of hypotonic medium) swelled to a spherical shape through an intermediate ellipsoidal form.

3. 3.At 49.5°C (where cells had cupped shapes prior to injection) the transition to the spherical form often involved a stomatocytic rather than ellipsoidal intermediate shape.

4. 4. The cupped form of the cells prior to injection did not account for the high incidence of cells swelling through a stomatocytic intermediate shape at 49.5°C.

5. 5. A theoretical treatment by Svetina and Zeks (1983) attributes the nature of the osmotic swelling transition shape to the difference in area between the outer and inner faces of the membrane. Our results would be consistent with the theoretical predictions if it is assumed that an increase in the area of the inner face of the membrane follows thermal denaturation of the cytoskeleton of cells in hypotonic medium.

The Mechanism of Isotonic Water Transport

The mechanism by which active solute transport causes water transport in isotonic proportions across epithelial membranes has been investigated. The principle of the experiments was to measure the osmolarity of the transported fluid when the osmolarity of the bathing solution was varied over an eightfold range by varying the NaCl concentration or by adding impermeant non-electrolytes. An in vitro preparation of rabbit gall bladder was suspended in moist oxygen without an outer bathing solution, and the pure transported fluid was collected as it dripped off the serosal surface. Under all conditions the transported fluid was found to approximate an NaCl solution isotonic to whatever bathing solution used. This finding means that the mechanism of isotonic water transport in the gall bladder is neither the double membrane effect nor co-diffusion but rather local osmosis. In other words, active NaCl transport maintains a locally high concentration of solute in some restricted space in the vicinity of the cell membrane, and water follows NaCl in response to this local osmotic gradient. An equation has been derived enabling one to calculate whether the passive water permeability of an organ is high enough to account for complete osmotic equilibration of actively transported solute. By application of this equation, water transport associated with active NaCl transport in the gall bladder cannot go through the channels for water flow under passive conditions, since these channels are grossly too impermeable. Furthermore, solute-linked water transport fails to produce the streaming potentials expected for water flow through these passive channels. Hence solute-linked water transport does not occur in the passive channels but instead involves special structures in the cell membrane, which remain to be identified.     

Senescence-Dependent Increase in the Permeability of Liposomes Prepared from Bean Cotyledon Membranes

Liposomes containing 79 mM Tris-acetate and 50 mM KCl were preparedfrom the total lipid extracts of smooth microsomal membranesisolated from 2, 4, 7 and 9 d old bean cotyledons. Permeabilityto glycerol was determined by spectrophotometric measurementsof osmotic swelling when the liposomes were placed in eitherisotonic or slightly hypotonic glycerol. For liposomes from2 and 4 d old membrane there was a time-dependent decrease inabsorbance at 450 nm from which initial swelling rates reflectingthe influx of glycerol and water were calculated. At 25 °Cthese rates were not significantly different For liposomes from7 and 9 d old membrane there was no change in absorbance withtime at 450 nm signifying that these older liposomes were equallypermeable and non-electrolytes and non-electrolytes, and thereforeincapable of swelling. Permeability to glucose was determinedby preparing the liposomes in a solution of the sugar, passingthem through a Sephadex column to eliminate unsequestered glucose,and quantifying sugar leaked from the liposomes over time bymeasuring NADPH formation through the tandem actions of hexokinaseand glucose-6-phosphate dehydrogenase. The rate constants forglucose leakage from 2 and 7 d old liposomes were 0.55 and 3.94respectively, again indicating a dramatic increase in permeabilitywith advancing age. These changes in permeability correlatetemporally with the appearance of gel phase lipid in both liposomesand the membranes from which they were derived, suggesting thatthe coexistence of discrete liquid-crystalline and gel phaselipid domains renders membranes leaky and contributes to lossof intracellular compartmentation in senescing tissue.     

Influence of homologous n-alkanoic on functional properties of isolated skeletal muscles. II. Membrane resting potential and the osmotic effectiveness of alkanoic acid]

The influence of butyric, hexanoic, octanoic, and decanoic acid on the membrane resting potential of isolated frog skeletal muscles were studied and the osmotic effects of n-alkanoic acids tested. 1. n-alkanoic acids cause osmotic effects like impermeable non-electrolytes (sucrose). Therefore, the permeability to alkanoic acids of the resting muscle cell membrane seems to be small. There are no differences between the acids tested. 2. The membrane resting potential is differently affected. Butyric acid in high concentration effects a hyperpolarization of the membrane whereas higher homologues (C6--C10) cause a depolarization. The depolarizing action increases with increasing concentration, exposure, and with the length of the hydrocarbon chain of the alkanoic acids. 3. It is suggested that osmotic effects are the cause for hyperpolarization of the membrane by high concentrations of butyric acid. 4. The depolarizing action of hexanoic, octanoic, and decanoic acid is discussed with regard to alterations induced by alkanoic acids in the membrane permeability and/or in the metabolism of the cells.     

The contributions of normal and anomalous osmosis to the osmotic effects arising across charged membranes with solutions of electrolytes

The osmotic effect arising across a porous membrane separating the solution of an electrolyte from water (or a more dilute solution) is ordinarily due to both normal osmosis, as it occurs also with non-electrolytes, and to "anomalous" osmosis. It is shown that the normal osmotic component cannot be measured quantitatively by the conventional comparison with a non-electrolytic reference solute. Anomalous osmosis does not occur with electroneutral membranes. Accordingly, with membranes which can be charged and discharged reversibly (without changes in geometrical structure), such as many proteinized membranes, the osmotic effects caused by an electrolyte can be measured both when only normal osmosis arises (with the membrane in the electroneutral state) and when normal as well as anomalous osmosis occurs (with the membrane in a charged state). The difference between these two effects is the true anomalous osmosis. Data are presented on the osmotic effects across an oxyhemoglobin membrane in the uncharged state at pH 6.75 and in two charged states, positive at pH 4.0 and negative at pH 10.0, with solutions of a variety of electrolytes using a concentration ratio of 2:1 over a wide range of concentrations. The rates of the movement of liquid across the membrane against an inconsequentially small hydrostatic head are recorded instead of, as conventional, the physiologically less significant pressure rises after a standard time.     

Mitochondria in the flight muscles of insects. I. Chemical composition and enzymatic content

1. The "indirect" thoracic muscles of adult dipterous and hymenopterous insects consist of a unique type of muscle characterized by the presence of numerous spherical, intracytoplasmic bodies termed "sarcosomes." 2. When the muscle is teased or ground, the sarcosomes are liberated as a turbid suspension of bodies ranging from 1 to 4 µ in diameter. A method is described for the isolation of sarcosomes by a simple differential centrifugation. 3. The cytochemical, chemical, and enzymatic properties of sarcosomes were examined for the purpose of appraising their relation to the cytoplasmic bodies of other tissues. 4. Fresh sarcosomes are slowly but selectively stained by the mitochondrial reagents, Janus green B and pinacyanol. Fixed sarcosomes give a positive reaction with Regaud's mitochondrial stain. 5. Chemical analyses show that approximately 29 per cent of the dry weight of sarcosomes consists of lipids and 60 per cent of protein. Microbiological assay indicates the presence of about 1 gamma of riboflavin per milligram of nitrogen. These values resemble those reported for isolated mitochondria of vertebrate liver and kidney. 6. When examined spectroscopically the sarcosomes, like the vertebrate mitochondria, show a high titer of cytochromes a, b, and c. 7. The titer of cytochrome oxidase varies systematically with the adult age of the insect. A similar relation is observed for the enzyme catalase. 8. Isolated sarcosomes show significant titers of succinoxidase, α-glycerophosphate dehydrogenase, malic dehydrogenase, and pyruvic dehydrogenase. The following dehydrogenases could not be demonstrated: xanthine, phenylalanine, glycine, lactic, choline, glutamic, and alcohol. These results are compared with those previously reported for vertebrate mitochondria. 9. In view of their manifold points of biochemical similarity, it is concluded that the sarcosomes are the mitochondria of this highly specialized muscular tissue.