A theory for the membrane potential of living cells

We give an explicit formula for the membrane potential of cells in terms of the intracellular and extracellular ionic concentrations, and derive equations for the ionic currents that flow through channels, exchangers and electrogenic pumps. We demonstrate that the work done by the pumps equals the change in potential energy of the cell, plus the energy lost in downhill ionic fluxes through the channels and exchangers. The theory is illustrated in a simple model of spontaneously active cells in the cardiac pacemaker. The model predicts the experimentally observed intracellular ionic concentration of potassium, calcium and sodium. Likewise, the shapes of the simulated action potential and five membrane currents are in good agreement with experiment. We do not see any drift in the values of the concentrations in a long time simulation, and we obtain the same asymptotic values when starting from the full equilibrium situation with equal intracellular and extracellular ionic concentrations. Received: 9 December 1998 / Revised version: 30 August 1999 / Accepted: 15 October 1999     

Osmotic tolerance limits and membrane permeability characteristics of stallion spermatozoa treated with cholesterol

Stallion spermatozoa exhibit osmotic damage during the cryopreservation process. Recent studies have shown that the addition of cholesterol to spermatozoal membranes increases the cryosurvival of bull, ram and stallion spermatozoa, but the exact mechanism by which added cholesterol improves cryosurvival is not understood. The objectives of this study were to determine if adding cholesterol to stallion sperm membranes alters the osmotic tolerance limits and membrane permeability characteristics of the spermatozoa. In experiment one, stallion spermatozoa were treated with cholesterol-loaded cyclodextrin (CLC), subjected to anisotonic solutions and spermatozoal motility analyzed. The spermatozoa were then returned to isotonic conditions and the percentages of motile spermatozoa again determined. CLC treatment increased the osmotic tolerance limit of stallion spermatozoa in anisotonic solutions and when returned to isotonic conditions. The second and third experiments utilized an electronic particle counter to determine the plasma membrane characteristics of stallion spermatozoa. In experiment two, stallion spermatozoa were determined to behave as linear osmometers. In experiment three, spermatozoa were treated with CLC, incubated with different cryoprotectants (glycerol, ethylene glycol or dimethyl formamide) and their volume excursions measured during cryoprotectant removal at 5° and 22 °C. Stallion spermatozoa were less permeable to the cryoprotectants at 5 °C than 22 °C. Glycerol was the least permeable cryoprotectant in control cells. The addition of CLC’s to spermatozoa increased the permeability of stallion spermatozoa to the cryoprotectants. Therefore, adding cholesterol to spermatozoal membranes reduces the amount of osmotic stress endured by stallion spermatozoa during cryopreservation.     

Osmotic tolerance and membrane permeability characteristics of rhesus monkey (Macaca mulatta) spermatozoa

Biophysical characteristics of the plasma membrane, such as osmotic sensitivity and water and cryoprotectant permeability are important determinants of the function of spermatozoa after cryopreservation. A series of experiments was conducted with rhesus macaque spermatozoa at 23 degrees C to determine their: (1) cell volume and osmotically inactive fraction of the cell volume; (2) permeability coefficients for water and the cryoprotectants dimethyl sulfoxide, glycerol, propylene glycol, and ethylene glycol; (3) tolerance to anisosmotic conditions; and (4) motility after a one step addition and removal of the four cryoprotectants. An electronic particle counter and computer aided semen analysis were used to determine the cell volume and permeability coefficients, and motility, respectively. Rhesus spermatozoa isosmotic cell volume was 27.7+/-3.0 microm3 (mean+/-SEM) with an osmotically inactive cell fraction of 51%. Hydraulic conductivity in the presence of dimethyl sulfoxide, glycerol, propylene glycol, and ethylene glycol was 1.09+/-0.30, 0.912+/-0.27, 1.53+/-0.53, and 1.94+/-0.47 microm/min/atm, respectively. Cryoprotectant permeability was 1.39+/-0.31, 2.21+/-0.32, 3.38+/-0.63, and 6.07+/-1.1 (x10(-3)cm/min), respectively. Rhesus sperm tolerated all hyposmotic exposures. However, greater than 70% motility loss was observed after exposure to solutions of 600 mOsm and higher. A one step addition and removal of all four cryoprotectants did not cause significant motility loss. These data suggest that rhesus sperm are tolerant to hyposmotic conditions, and ethylene glycol may be the most appropriate cryoprotectant for rhesus sperm cryopreservation, as it has the highest permeability coefficient of the tested cryoprotectants.     

Osmotic fragility and fluidity of erythrocyte membranes from rats raised on an essential fatty acid deficient diet A spin label study

Erythrocyte membranes from rats raised on a diet with low content of essential fatty acids were studied by osmotic sensitivity tests and spin labeling techniques. This diet induced significant modifications in acylglycerophosphocholine fatty acid composition with regard to 16 : 1, 18 : 1, 18 : 2 (n-6), 20 : 3 (n-9), and 20 : 4 (n-6). No changes in membrane fluidity as monitored by spin label motion were found but the diet caused an increased osmotic sensitivity in essential fatty acid deficient erythrocytes. 50% hemolysis was obtained at a 51.0% dilution of saline with H₂O as compared to a 57.0% dilution for the control material. Membrane fluidity was unaffected by γ-irradiation up to 80 krad.     

Osmotic tolerance limits and effects of cryoprotectants on the motility, plasma membrane integrity and acrosomal integrity of rat sperm

Osmotic stress is an important factor that can result in cell damage during cryopreservation. The objectives of this study were to determine: (1) isosmotic sperm cell volume; (2) osmotically inactive volume; (3) osmotic tolerance limits of rat sperm; and (4) the effects of addition and removal of glycerol (Gly), ethylene glycol (EG), propylene glycol (PG) or dimethyl sulfoxide (Me(2)SO) on rat sperm function. Sperm from Fischer 344 and Sprague-Dawley rats were used in this study. An electronic particle counter was used to measure the cell volume of rat sperm. Computer-assisted sperm motility analysis and flow-cytometric analysis were used to assess sperm motility, plasma membrane and acrosomal integrity. The isosmotic sperm cell volumes of the two strains were 37.0+/-0.1 and 36.2+/-0.2 microm(3), respectively. Rat sperm behaved as linear osmometers from 260 to 450 mOsm, and the osmotically inactive sperm volumes of the two strains were 79.8+/-1.5% and 81.4+/-2.2%, respectively. Rat sperm have very limited osmotic tolerances. The sperm motility and the sperm plasma membranes of both strains were sensitive to anisosmotic treatments, but the acrosomes of both strains were more sensitive to hyposmotic than hyperosmotic conditions. The one-step addition and removal of Me(2)SO showed the most deleterious effect on rat sperm motility, plasma membrane integrity, and acrosomal integrity among the four cryoprotectants. These data characterizing rat sperm osmotic behavior, osmotic and cryoprotectant tolerance will be used to design cryopreservation protocols for rat sperm.     

A secondary disruption of the dmpA gene encoding a large membrane protein allows aggregation defective Dictyostelium rasC- cells to form multicellular structures

The disruption of the gene encoding the Dictyostelium Ras subfamily protein, RasC, results in a strain that does not aggregate and has defects in both cAMP signal relay and cAMP chemotaxis. Disruption of a second gene in the rasC(-) strain by Restriction Enzyme Mediated Integration produced cells that were capable of forming multicellular structures in plaques on bacterial lawns. The disrupted gene (dmpA) encoded a novel membrane protein that was designated Dmp1. Although the rasC(-)/dmpA(-) cells progressed through early development, they did not form aggregation streams on a plastic surface under submerged starvation conditions. Phosphorylation of PKB in response to cAMP, which is significantly reduced in rasC(-) cells, remained low in the rasC(-)/dmpA(-) cells. However, in spite of this low PKB phosphorylation, the rasC(-)/dmpA(-) cells underwent efficient chemotaxis to cAMP in a spatial gradient. Cyclic AMP accumulation, which was greatly reduced in the rasC(-) cells, was restored in the rasC(-)/dmpA(-) strain, but cAMP relay in these cells was not apparent. These data indicate that although the rasC(-)/dmpA(-) cells were capable of associating to form multicellular structures, normal aggregative cell signaling was clearly not restored. Disruption of the dmpA gene in a wild-type background resulted in cells that exhibited a slight defect in aggregation and a more substantial defect in late development. These results indicate that, in addition to the role played by Dmp1 in aggregation, it is also involved in late development.     

Osmotic tolerance of avian spermatozoa: influence of time, temperature, cryoprotectant and membrane ion pump function on sperm viability

Potential factors influencing sperm survival under hypertonic conditions were evaluated in the Sandhill crane (Grus canadensis) and turkey (Meleagridis gallopavo). Sperm osmotolerance (300-3000 mOsm/kg) was evaluated after: (1) equilibration times of 2, 10, 45 and 60 min at 4 °C versus 21 °C; (2) pre-equilibrating with dimethylacetamide (DMA) or dimethylsulfoxide (Me₂SO) at either 4 °C or 21 °C; and (3) inhibition of the Na⁺/K⁺ and the Na⁺/H⁺ antiporter membrane ionic pumps. Sperm viability was assessed using the eosin-nigrosin live/dead stain. Species-specific differences occurred in response to hypertonic conditions with crane sperm remaining viable under extreme hypertonicity (3000 mOsm/kg), whereas turkey sperm viability was compromised with only slightly hypertonic (500 mOsm/kg) conditions. The timing of spermolysis under hypertonic conditions was also species-specific, with a shorter interval for turkey (2 min) than crane (10 min) sperm. Turkey sperm osmotolerance was slightly improved by lowering the incubation temperature from 21 to 4 °C. Pre-equilibrating sperm with DMA reduced the incidence of hypertonic spermolysis only in the crane, at both room and refrigeration temperature. Inhibiting the Na⁺/K⁺ and the Na⁺/H⁺ antiporter membrane ion pumps did not impair resistance of crane and turkey spermatozoa to hypertonic stress; pump inhibition actually increased turkey sperm survival compared to control sperm. Results demonstrate marked species specificity in osmotolerance between crane and turkey sperm, as well as in the way temperature and time of exposure affect sperm survival under hypertonic conditions. Differences are independent of the role of osmotic pumps in these species.     

Proton-induced endocytosis is dependent on cell membrane fluidity,lipid-phase order and the membrane resting potential

Recently it has been shown that decreasing the extracellular pH of cells stimulates the formation of inward membrane invaginations and vesicles, accompanied by an enhanced uptake of macromolecules. This type of endocytosis was coined as proton-induced uptake (PIU). Though the initial induction of inward membrane curvature was rationalized in terms of proton-based increase of charge asymmetry across the membrane, the dependence of the phenomenon on plasma membrane characteristics is still unknown. The present study shows that depolarization of the membrane resting potential elevates PIU by 25%, while hyperpolarization attenuates it by 25%. Comparison of uptake in suspended and adherent cells implicates that the resting-potential affects PIU through remodeling the actin-cytoskeleton. The pH at the external interface of the cell membrane rather than the pH gradient across it determines the extent of PIU. PIU increases linearly upon temperature increase in the range of 4–36 °C, in correlation with the membrane fluidity. The plasma membrane fluidity and the lipid phase order are modulated by enriching the cell's membrane with cholesterol, tergitol, dimethylsulfoxide, 6-ketocholestanol and phloretin and by cholesterol depletion. These treatments are shown to alter the extent of PIU and are better correlated with membrane fluidity than with the lipid phase order. We suggest that the lipid phase order and fluidity influence PIU by regulating the lipid order gradient across the perimeter of the lipid-condensed microdomains (rafts) and alter the characteristic tension line that separates the higher ordered lipid-domains from the lesser ordered ones.     

Osmotic tolerance of mouse spermatozoa from various genetic backgrounds: acrosome integrity, membrane integrity, and maintenance of motility

All cells have an intrinsic biophysical property related to their ability to undergo osmotically driven volume changes. This project is of fundamental importance to our understanding of the basic cryobiology of mouse spermatozoa. The objectives of this study were to determine the osmotic tolerance limits for (1) motility, (2) acrosome integrity, and (3) membrane integrity of mouse spermatozoa from multiple genetic backgrounds including: C57BL/6, BALB/c, FVB, C3H, 129/SVS2 hsd B6C3F1, CB6F1, and ICR. The maintenance of acrosomal and plasma membrane integrity was not affected by genetic background (p=0.13), however, there was an interaction between genetic background and osmolality. In addition, acrosome and plasma membrane integrity was highly correlated within each strain (p<0.01). In contrast to acrosome and plasma membrane integrity, the motility of spermatozoa from different genetic backgrounds fell sharply on both sides of isosmolality, both with and without return to isosmotic conditions. Exposure to hyposmotic conditions caused morphological changes in the spermatozoa, which inhibited motility. However, this morphological change was not reversible in all cases when returned to isosmotic conditions. The ability to maintain motility in an anisosmotic media was affected by genetic background, osmolality as well as the interaction between genetic background and osmolality (p<0.05). In conclusion, mice with different genetic backgrounds appear to have similar tolerance to osmotic changes in terms of sperm acrosome and plasma membrane integrity; however, the ability to maintain motility differs between genetic backgrounds.     

Role of membrane thermotropic properties on hypotonic hemolysis and hypertonic cryohemolysis of human red blood cells

The hypothesis of a correlation between the effects of temperature on red blood cells hypotonic hemolysis and hypertonic cryohemolysis and two thermotropic structural transitions evidenced by EPR studies has been tested. Hypertonic cryohemolysis of red blood cells shows critical temperatures at 7 degrees C and 19 degrees C. In hypotonic solution, the osmotic resistance increases near 10 degrees C and levels off above 20 degrees C. EPR studies of red blood cell membrane of a 16-dinyloxyl stearic acid spin label show, in the 0-50 degrees C range, the presence of three thermotropic transitions at 8, 20, and 40 degrees C. Treatments of red blood cells with acidic or alkaline pH, glutaraldehyde, and chlorpromazine abolish hypertonic cryohemolysis and reduce the effect of temperature on hypotonic hemolysis. 16-Dinyloxyl stearic acid spectra of red blood cells treated with glutaraldehyde and chlorpromazine show the disappearance of the 8 degrees C transition. Both the 8 degrees C and the 20 degrees C transitions were abolished by acidic pH treatment. The correlation between the temperature dependence of red blood cell lysis and thermotropic breaks might be indicative of the presence of structural transitions producing areas of mismatching between differently ordered membrane components where the osmotic resistance is decreased.     

Efficient mechanical disruption of Lactobacillus helveticus,Lactococcus lactis and Propionibacterium freudenreichii by a new high-pressure homogenizer and recovery of intracellular aminotransferase activity

Microbiological studies often involve bacterial cell fractionation, which is known to be difficult for Gram-positive as compared to Gram-negative bacteria. Our purpose was to test the breaking efficiency of a new high-pressure pilot homogenizer for three Gram-positive species involved in dairy technology and to assess the activity of an intracellular aminotransferase. Varied pressures (50, 100 and 200 MPa) were applied to concentrated bacterial suspensions (1.2 mg dry weight/ml) of Lactobacillus helveticus, Lactococcus lactis and Propionibacterium freudenreichii. Breaking efficiency was estimated by decreases in optical density at 650 nm, cellular dry weight and viability. The proteins released were quantified and the residual intracellular aminotransferase activity was estimated using leucine as substrate. One run at 50 MPa was sufficient to break 80% of lactobacilli cells whereas 200 MPa were required for the same efficiency for L. lactis and P. freudenreichii. Whatever the pressure, leucine aminotransferase activity was recovered in the supernatant after cell breaking. This new high-pressure pilot homogenizer can allow rapid (20 s/run), easy, continuous and highly efficient cell breaking for intracellular enzyme recovery or other purposes. As the species tested were not phylogenetically related, and had different morphologies and cell wall compositions, we conclude that most Gram-positive bacteria may be broken efficiently by this new device. Electronic Publication     

Sequence and temperature effect on hydrogen bond disruption in DNA determined by a statistical analysis

We use the modified self-consistent phonon approximation theory to calculate temperature dependent interbase hydrogen bond disruption profiles for a number of six base pair repeating sequence infinite B-DNA polymers with various guanine-cytosine/adenine-thymine ratios. For comparison we also include results we have obtained in our earlier work on several B-DNA homopolymers, copolymers and a four-base-pair repeating sequence polymer. Our theory gives a statistical estimate of thermal fluctuational disruption probability of individual hydrogen bonds in individual base pairs in DNA as a function of temperature. The calculated probabilities show no sequence dependence at premelting temperatures, in agreement with proton exchange measurements. These probabilities however become very sensitive to base sequence at temperatures close to the observed melting temperatures. Multi-phasic critical transitions are found in which a portion of base pairs are disrupted at temperatures below the final disruption temperature. These transitions include localized as well as non-localized base pair opening. The localized transitions involve disruption of a few base-pairs at every other location without large scale base unstacking, and they may not appear in the observed UV curves with current resolution. On the other hand the overall disruption behavior is consistent with observations. The midpoint transition temperatures are close to the observed melting temperatures and these temperatures show the observed linear dependence on guanine-cytosine content. Our calculations indicate that our theory can be used effectively to calculate H-bond disruption behavior of different DNA sequences. Received: 20 February 1996 / Accepted: 2 May 1996     

Mouse myodulin, a new potential angiogenic factor, functionally expressed in yeast

Myodulin is a new integral membrane protein down-regulated in skeletal muscle atrophy. A first characterization suggested that myodulin could be a skeletal muscle angiogenic factor operating through direct cell-to-cell interactions. Here, we show that mouse myodulin can be expressed at the plasma membrane of Saccharomyces cerevisiae and purified. Co-culture experiments of myoblasts and cardiac vascular endothelial cells reveal that myodulin, either presented in yeast membranes or in liposomes after purification, increases the invasive potential of endothelial cells with a similar efficiency as when over-expressed in skeletal muscle cells. Functional essays using myodulin expressed in yeast bring new information about the myodulin functional mechanism, suggesting that one or several muscle cell components could be necessary for myodulin to increase the invasive potential of endothelial cells. The yield of purified myodulin should allow structure-function relationships studies for a better understanding of myodulin functional mechanisms.     

Interactions of oritavancin, a new lipoglycopeptide derived from vancomycin, with phospholipid bilayers: Effect on membrane permeability and nanoscale lipid membrane organization

Antibiotics acting on bacterial membranes are receiving increasing attention because of widespread resistance to agents acting on other targets and of potentially improved bactericidal effects. Oritavancin is a amphiphilic derivative of vancomycin showing fast and extensive killing activities against multi-resistant (including vancomycin insusceptible) Gram-positive organisms with no marked toxicity towards eukaryotic cells. We have undertaken to characterize the interactions of oritavancin with phospholipid bilayers, using liposomes (LUV) and supported bilayers made of cardiolipin (CL) or phosphatidylglycerol (POPG) and phosphatidylethanolamine (POPE), all abundant in Gram-positive organisms. Changes in membrane permeability were followed by the release of calcein entrapped in liposomes at self-quenching concentrations, and changes in nanoscale lipid organization examined by Atomic Force Microscopy (AFM). Oritavancin caused a fast (< 5 min) and complete (> 95%) release of calcein from CL:POPE liposomes, and a slower but still substantial (50% in 60 min) release from POPG:POPE liposomes, which was (i) concentration-dependent (0-600 nM; [microbiologically meaningful concentrations]); (ii) enhanced by an increase in POPG:POPE ratio, and decreased when replacing POPG by DPPG. AFM of CL:POPE supported bilayers showed that oritavancin (84 nM) caused a remodeling of the lipid domains combined with a redisposition of the drug and degradation of the borders. In all the above studies, vancomycin was without a significant effect at 5.5 μM. Electrostatic interactions, together with lipid curvature, lipid polymorphism as well of fluidity play a critical role for the permeabilization of lipid bilayer and changes in lipid organization induced by oritavancin.     

A stochastic model for the membrane potential of a stimulated neuron

We present a simple model describing the transition between the prefiring, firing and postfiring phases of a single neuron in a large neural net. Using typical values for the physiological parameters that enter the model, we find average interspike times that are close to those reported in experimental measurements.     

Determination of oocyte membrane permeability coefficients and their application to cryopreservation in a rabbit model

Having an effective means to cryopreserve human oocytes would offer more flexibility in healthcare services for infertility patients, and obviate cryopreservation of preimplantation embryos. It is essential to establish good animal models for human oocyte cryopreservation and the rabbit is a good candidate. Attempts to improve oocyte cryopreservation are often empirical, with results often being irreproducible. Cryopreservation protocols may be optimized by modeling the changes in oocyte volume and the associated damages incurred during the addition and dilution of cryoprotective agents (CPA). The objectives of the current study were to determine cryobiological properties of rabbit oocytes, including the isotonic volume, osmotically inactive cell fraction (V_b), hydraulic conductivity (L_p), permeability (P_s) to dimethylsulfoxide (Me₂SO), ethylene glycol (EG), and glycerol (GLY) and to examine the correlation between cell volume excursions and viability. This has led to the development of the accumulative osmotic damage (AOD) model associated with the processes of CPA addition/dilution. Mature rabbit oocytes were perfused with 15% (V/V) CPA medium (dissolved in 1× PBS). The osmotic responses of the oocytes were videotaped. A two-parameter model was fit to the experimental data to determine the values of L_p and P_s. Oocyte volumes reached upon equilibration with 285, 600, 900, and 1200 mOsm (milliosmolal) solutions of non-permeating compounds were plotted in a Boyle van’t Hoff plot. The average radius of rabbit oocytes in an isotonic solution was determined to be 55.7 ± 1.2 μm (n = 16). The rabbit oocyte exhibited an “ideal” osmotic response in the range from iso-osmolity to 1200 mOsm. The V_b was determined to be 20% of the isotonic value with r² = 0.97. The values of L_p were determined to be 0.79 ± 0.26, 0.82 ± 0.22, and 0.64 ± 0.16 μm min⁻¹ atm⁻¹ and the P_s values were determined to be 2.9 ± 1.3, 2.7 ± 1.3, and 0.27 ± 0.18 × 10⁻³ cm min⁻¹ for Me₂SO, EG and GLY, respectively. There were no significant differences (p > 0.05) between values for L_p and P_S in the presence of the Me₂SO and EG. However, these values were significantly different from the values in presence of GLY. We calculated the AOD values of those oocytes that experienced the process of CPA additions/dilutions and found that these values were highly correlated to the development rates of these oocytes after parthenogenetic activation (r = −0.98).     

A model of plasma membrane flow and cytosis regulation in growing pollen tubes

A model of cytosis regulation in growing pollen tubes is developed and simulations presented. The authors address the question on the minimal assumptions needed to describe the pattern of exocytosis and endocytosis reported recently by experimental biologists. Biological implications of the model are also treated. Concepts of flow and conservation of membrane material are used to pose an equation system, which describes the movement of plasma membrane in the tip of growing pollen tubes. After obtaining the central equations, relations describing the rates of endocytosis and exocytosis are proposed. Two cytosis receptors (for exocytosis and endocytosis), which have different recycling rates and activation times, suffice to describe a stable growing tube. Simulations show a very good spatial separation between endocytosis and exocytosis, in which separation is shown to depend strongly on exocytic vesicle delivery. In accordance to measurements, most vesicles in the clear zone are predicted to be endocytic. Membrane flow is essential to maintain cell polarity, and bi-directional flow seems to be a natural consequence of the proposed mechanism. For the first time, a model addressing plasma membrane flow and cytosis regulation were posed. Therefore, it represents a missing piece in an integrative model of pollen tube growth, in which cell wall mechanics, hydrodynamic fluxes and regulation mechanisms are combined.     

Effect of mechanical abrasion on the viability, disruption and germination of spores of Bacillus subtilis

AIMS: To elucidate the factors influencing the sensitivity of Bacillus subtilis spores in killing and disrupting by mechanical abrasion, and the mechanism of stimulation of spore germination by abrasion. METHODS AND RESULTS: Spores of B. subtilis strains were abraded by shaking with glass beads in liquid or the dry state, and spore killing, disruption and germination were determined. Dormant spores were more resistant to killing and disruption by abrasion than were growing cells or germinated spores. However, dormant spores of the wild-type strain with or without most coat proteins removed, spores of strains with mutations causing spore coat defects, spores lacking their large depot of dipicolinic acid (DPA) and spores with defects in the germination process exhibited essentially identical rates of killing and disruption by abrasion. When spores lacking all nutrient germinant receptors were enumerated by plating directly on nutrient medium, abrasion increased the plating efficiency of these spores before killing them. Spores lacking all nutrient receptors and either of the two redundant cortex-lytic enzymes behaved similarly in this regard, but the plating efficiency of spores lacking both cortex-lytic enzymes was not stimulated by abrasion. CONCLUSIONS: Dormant spores are more resistant to killing and disruption by abrasion than are growing cells or germinated spores, and neither the complete coats nor DPA are important in spore resistance to such treatments. Germination is not essential for spore killing by abrasion, although abrasion can trigger spore germination by activation of either of the spore's cortex-lytic enzymes. SIGNIFICANCE AND IMPACT OF THE STUDY: This work provides new insight into the mechanisms of the killing, disruption and germination of spores by abrasion and makes the surprising finding that at least much of the spore coat is not important in spore resistance to abrasion.