Biomedical subjects

W C Ford

Publications and source records attributed to W C Ford.

At least 55 records · Page 3Linked to original sources

PubMed1990

A comparison of the freezing of human semen in the uncirculated vapour above liquid nitrogen and in a commercial semi-programmable freezer.

The cooling rates inside 0.25-ml semen straws filled with glycerol, egg yolk, citrate buffer were compared between a standard vapour freezing procedure and freezing in a Nicool LM-10 semi-programmable freezer. During vapour freezing, the cooling rate at the bottom of the straw was much faster than at the top and there was considerable variation between replicates. By contrast, position within the straw did not affect the cooling rate in the Nicool LM-10 procedure and replicates were more consistent. More motile spermatozoa survived the Nicool freezing procedure and their lateral head displacement was greater than vapour frozen spermatozoa. However the percentage of intact spermatozoa and their velocity was very similar after the two procedures. We conclude that the freezing procedure using the Nicool LM-10 provides a comparatively economical way to achieve consistent semen freezing for research studies.

Cell Membrane↗

PubMed1990

Effect of caffeine and of pentoxifylline on the motility and metabolism of human spermatozoa.

Human spermatozoa were washed and incubated with 6 mM-caffeine or 0.15-1.2 mM-pentoxifylline. Sperm motility was measured by time-lapse photography, the rate of glycolysis by the release of tritiated water from 1 mM-[3-3H]D-glucose and the rate of mitochondrial respiration by the release of 14CO2 from 1 mM-[U-14C]-L-lactate or 1 mM-[2-14C]pyruvate. Caffeine stimulated the majority of spermatozoa to convert from the 'rolling' to the 'yawing' mode of progression with a concomitant increase in lateral head displacement from 4.1 +/- 0.09 microns (343) to 6.7 +/- 0.25 microns (105) (mean +/- s.e.m. (number of spermatozoa)). There was a 45% decline in the percentage of progressively motile spermatozoa and a very small decrease in their velocity. Pentoxifylline had only a slight effect on lateral head displacement or percentage motility but produced a significant increase in velocity. Both compounds increased the rate of glycolysis by greater than 40% but elevated the rate of 14CO2 production to a smaller extent. The concentrations of ATP and ADP changed very little. We conclude that the glycolytic pathway in human spermatozoa can respond efficiently to changes in energy demand.

Adenosine Diphosphate↗

PubMed1988

An evaluation of the role of relaxin in the penetration of cervical mucus by spermatozoa.

Relaxin-like immunoreactivity was measured in seminal plasma from men who were separated into two groups, on the basis of a previous positive or negative result in a postcoital cervical mucus penetration test. There was no difference in the relaxin concentration between the groups. The effect of exogenous porcine relaxin (0, 10 or 100 ng/ml) on human cervical mucus penetration in vitro by washed human spermatozoa was studied using a capillary tube preparation. In the positive postcoital test group the highest relaxin concentration (100 ng/ml) tended to inhibit cervical mucus penetration, although this effect was only significant for one of the parameters measured (number of spermatozoa penetrating to the 10-mm mark). The same trend was apparent for the negative postcoital test group, but no differences were significant. The results are in direct contrast to previous reports that relaxin can stimulate human spermatozoa motility and cervical mucus penetration.

Cervix Mucus↗

PubMed1987

Futile substrate cycles in the glycolytic pathway of boar and rat spermatozoa and the effect of alpha-chlorohydrin.

In boar spermatozoa incubated with 0.1 mM-glucose about 20 nmol glucose were converted to lactate and CO2 and the rate of futile substrate cycling between glucose and glucose 6-phosphate was about 6 nmol/10(8) spermatozoa/30 min. Futile cycling was increased in the presence of 0.05 or 1 mM-alpha-chlorohydrin but not to an extent sufficient to account for the rapid decline in ATP concentration observed under these conditions. These estimates include a substantial rate of fructose formation from fructose phosphates. The addition of 10 mM-L-lactate plus 1 mM-pyruvate protected the spermatozoa against the effect of alpha-chlorohydrin and glucose on the ATP concentration but increased futile substrate cycling. Substrate cycling between fructose 6-phosphate and fructose 1,6-bisphosphate could not be measured in boar spermatozoa but in rat spermatozoa its rate (nmol/10(8) spermatozoa/30 min) was about 10 under control condition and about 25 in the presence of 1 mM-alpha-chlorohydrin. This increase was insufficient to account for the decline in ATP concentration. In both species futile substrate cycling consumed a significant proportion of the ATP synthesis during lactate production but only about 5% of that produced in the oxidation of glucose to acetyl carnitine and CO2.

Adenosine Triphosphate↗

PubMed1986

The concerted effect of alpha-chlorohydrin and glucose on the ATP concentration in spermatozoa is associated with the accumulation of glycolytic intermediates.

In the absence of a glycolysable sugar the effect of 1 mM-RS-alpha-chlorohydrin on the ATP concentration in ram or boar spermatozoa was relatively small but the addition of 0.10 or 0.03 mM-glucose initiated a rapid loss of ATP. When the spermatozoa were incubated with 0.05 mM-RS-alpha-chlorohydrin, the addition of 1.0 mM (ram) or 0.06 mM (boar)-glucose was required to produce ATP dissipation. In ram spermatozoa treated with 0.05 or 1.00 mM-RS-alpha-chlorohydrin, ATP loss was caused by 10 mM-fructose or 10 mM-mannose but not by 10 mM-glycerol or 10 mM-inositol. In boar spermatozoa incubated with 1 mM-RS-alpha-chlorohydrin the addition of 10 mM-L-lactate plus 1.0 mM-pyruvate protected the spermatozoa against the ability of 1.0 mM-glucose to produce a decline in ATP concentration. Every combination of treatments capable of inducing a marked decline in ATP concentration also caused a dramatic (20-100-fold) increase in the concentration of fructose 1,6-bisphosphate. An increase in fructose 1,6-bisphosphate concentration was never observed when the ATP concentration was unaffected. We conclude that it is very probable that the concerted effect of alpha-chlorohydrin and glycolysable sugar is responsible for the contraceptive action of alpha-chlorohydrin in vivo and that fructose 1,6-bisphosphate is implicated in its mechanism of action.

Adenosine Triphosphate↗

PubMed1985

The presence of glucose increases the lethal effect of alpha-chlorohydrin on ram and boar spermatozoa in vitro.

Ram cauda epididymal spermatozoa were incubated for 10 min at 34 degrees C with or without 1.0 mM-RS-alpha-chlorohydrin before (1) 5 mM-D-glucose or (2) 10 mM-L-lactate plus 1 mM pyruvate or (3) 5 mM-D-glucose plus 10 mM-L-lactate plus 1 mM-pyruvate or (4) no substrate was added. Without alpha-chlorohydrin, the motility, the ATP concentration and the energy charge of the spermatozoa were maintained for 240 min by substrate combinations 1-3 but with no added substrate (4) the motility declined after 60 min. All the values decreased dramatically after 10 min in spermatozoa exposed to alpha-chlorohydrin in substrate conditions 1 and 3 (glucose present) but alpha-chlorohydrin had no significant effect in conditions 2 and 4 (no glucose) except after prolonged incubation. In a dose-response experiment glucose-dependent ATP dissipation began to occur with 0.025 mM-RS-alpha-chlorohydrin. A similar effect was seen in boar spermatozoa exposed to 0.1-5.0 mM-alpha-chlorohydrin and 5 mM-D-glucose. With boar spermatozoa the presence of 10 mM-L-lactate and 1 mM-pyruvate as well as glucose prevented the loss of ATP. We conclude that this concerted action of alpha-chlorohydrin and glucose is probably responsible for the contraceptive action of alpha-chlorohydrin and propose that it may depend on 'futile substrate cycling' in the glycolytic pathway.

Adenosine Triphosphate↗

PubMed1984

The role of citrate in determining the activity of calcium ions in human semen.

The concentrations of calcium and citrate in 52 samples of human semen obtained from 10 donors were 11.07 +/- 0.41 (mean +/- SEM) and 27.2 +/- 1.62 mM, respectively, and were strongly correlated (r = + 0.79). The pH of the semen was 7.65 +/- 0.069. All 3 parameters varied significantly between the donors (P less than 0.001 in each case), but the activity of calcium ions measured with a calcium electrode was 0.24 +/- 0.022 mM (n = 45) and did not vary significantly between donors. The expected activity of calcium ions was calculated on the hypothesis that the chelation of calcium by citrate could explain the discrepancy between the concentration and the activity of the ion. The result was 0.57 +/- 0.043 mM (n = 52). It was concluded that chelation by citrate could be the most important factor in determining the low activity of Ca++ in semen.

Calcium↗

PubMed1984

The effect of experimentally induced diabetes on the metabolism of glucose by seminiferous tubules and epididymal spermatozoa from the rat.

The concentration of glucose in the plasma of alloxan-diabetic rats was 23.4 +/- 0.86 mM (mean +/- SEM; n = 18), and the concentration of insulin was 11.4 +/- 1.67 microU/ml (mean +/- SEM; n = 17). The weights of the ventral prostate (0.45 +/- 0.03 vs. 0.72 +/- 0.04 g) and seminal vesicles (1.23 +/- 0.06 vs. 1.84 +/- 0.08 g) were decreased compared to control values and the rats lost body weight, but the weights of the testes were not significantly different from control values (3.14 +/- 0.08 vs. 3.23 +/- 0.14 g/pair). Similar changes were seen in streptozotocin-diabetic rats. The concentration of fructose (micromoles per g fresh wt) was greater in the coagulating gland of alloxan-diabetic (19.6 +/- 1.3; n = 17) than control rats (9.1 +/- 0.7; n = 18). The production of 14CO2 from D-[U-14C]glucose by spermatozoa or seminiferous tubules from diabetic rats was decreased compared to that in controls [28 +/- 3 vs. 53 +/- 6 nmol glucose converted/10(8) spermatozoa X 30 min (n = 8) and 0.81 +/- 0.03 vs. 1.08 +/- 0.03 mumol glucose converted/g fresh wt X 30 min (n = 7)]. There was no change in the production of lactate or 3HOH from D-[2-3H] glucose, and the presence of insulin (10 mU/ml) in the incubation had little effect. Rat epididymal spermatozoa took up 2-deoxy-D-glucose by a facilitated diffusion mechanism; the Km was about 0.2 mM, with a maximum velocity of about 0.10 nmol/10(6) spermatozoa X 10 sec. Neither alloxan-diabetes nor the presence of insulin (10 mU/ml) had an appreciable effect on these parameters.

Alloxan↗

PubMed1983

The activity of glyceraldehyde 3-phosphate dehydrogenase in spermatozoa from different regions of the epididymis in laboratory rodents treated with alpha-chlorohydrin or 6-chloro-deoxyglucose.

The activity of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (mUnits/10(6) spermatozoa: mean +/- s.e.m., N = 12) in spermatozoa from the rat epididymis declined from 22.0 +/- 1.4 in the caput to 14.1 +/- 1.3 in the corpus region but there was no further decrease in the cauda region. In hamsters (N = 4), GAPDH activity in spermatozoa declined from 24.8 +/- 2.2 in the caput to 15.2 +/- 1.2 in the distal cauda epididymidis with the greatest decrease between the corpus and proximal cauda regions. In guinea-pigs (N = 4) GAPDH activity in spermatozoa increased from 11.4 +/- 0.79 in the caput to 18.0 +/- 0.7 in the corpus and cauda regions of the epididymis. The activity of GAPDH in spermatozoa therefore changes during maturation in a species dependent manner. GAPDH in spermatozoa from the distal cauda epididymidis of rats given alpha-chlorohydrin (4, 8 or 25 mg/kg/day by mouth) or 6-chloro-6-deoxyglucose (24 or 96 mg/kg/day by mouth) for 10 days was inhibited by greater than 80% but was only inhibited by 25-45% in spermatozoa from the caput epididymidis. The enzyme was inhibited to an intermediate and dose-dependent extent in spermatozoa from the corpus region. A similar pattern of inhibition was seen in spermatozoa from hamsters given alpha-chlorohydrin (50 or 100 mg/kg/day) for 10 days. alpha-Chlorohydrin (66 mg/kg/day s.c.) for 10 days inhibited GAPDH in spermatozoa from the caput or corpus epididymidis of the guinea-pig by less than 20% but decreased GAPDH activity by almost 90% in the cauda region. In rats the greater effect of alpha-chlorohydrin on spermatozoa from the cauda region of the epididymis occurred even after short periods of treatment or when the passage of spermatozoa through the duct was interrupted by a ligature around the corpus region, indicating that the effect is not simply a reflection of the length of time the spermatozoa have spent in the epididymis. It is concluded that either spermatozoa undergo a maturational change which increases their sensitivity to alpha-chlorohydrin or that alpha-chlorohydrin (or an active metabolite) is concentrated in the lumen of the cauda epididymidis.

Animals↗

PubMed1983

D-[1-14C]mannitol and [U-14C]sucrose as extracellular space markers for human spermatozoa and the uptake of 2-deoxyglucose.

[U-14C]Sucrose and D-[1-14C]mannitol were used to determine the tritiated water space of human spermatozoa to validate these compounds as markers for the extracellular space. Calculations based on 0.03 mM-[U-14C]sucrose gave a negative water space. The water space estimated with 0.03 mM-D[1-14C]mannitol was unstable but a stable result was obtained with 0.3 mM-D-[1-14C]mannitol in incubations up to 2 h. The mean water space was 2.21 +/- 0.106 microliters/10(8) spermatozoa (mean +/- s.e.m. for 6 batches of pooled semen). The water space was decreased or abolished by Triton X-100, cold shock, sonication or hypotonic treatment. The water space responded to changes in the osmolarity of the medium by increasing in dilute media. It is concluded that mannitol is an effective extracellular marker for human spermatozoa if concentrations greater than or equal to 0.3 mM are used. When the kinetics of the uptake of 2-deoxyglucose by the spermatozoa were studied by using mannitol as an extracellular marker, the transport was saturable and was inhibited by cytochalasin B. The Km was 1.6 +/- 0.33 mM and the Vmax was 4.2 +/- 0.52 nmol/10(8) spermatozoa/10 sec (mean +/- s.e.m., n = 4).

Biological Transport↗

PubMed1982

The effect of 6-deoxy-6-fluoroglucose on the fertility of male rats and mice.

A single dose of 6-deoxy-6-fluoroglucose (120 mg/kg) inhibited spermatogenesis in a group of 6 rats. The rats became infertile 16 to 30 days after the dose but 5 had recovered after 45 days. Both testes of the rat which remained infertile and one testis in each of 3 other rats had hypotrophied and damaged tubules were present even in the testes which remained of normal size. 6-Deoxy-6-fluoroglucose (0, 240, 480 or 960 mg/kg) did not affect the fertility of male mice. 10 mM-6-Deoxy-6-fluoroglucose caused a significant inhibition of glycolysis in rat seminiferous tubules in vitro but lower concentrations did not. There was no effect on the citrate concentration in the tubules.

Animals↗

PubMed1982

Activities of various 6-chloro-6-deoxysugars and (S) alpha-chlorohydrin in producing spermatocoeles in rats and paralysis in mice and in inhibiting glucose metabolism in bull spermatozoa in vitro.

6-Chloro-6-deoxyglucose, 6-chloro-6-deoxymannose, 6-chloro-6-deoxy-fructose, 6-chloro-6-deoxyglucitol, 6-chloro-6-deoxygalactose and (S) alpha-chlorohydrin all produced spermatocoeles in the ductuli efferentes and epididymis of the rat and were neurotoxic in the mouse, but only alpha-chlorohydrin caused substantial inhibition of glucose metabolism in bull spermatozoa in vitro. The relative potencies of the compounds in producing spermatocoeles reflected their activities as reversible antifertility agents in the rat but compared to the others 6-chloro-6-deoxymannose was considerably less neurotoxic to mice than might have been anticipated from its contraceptive dose. Thus different metabolites may be responsible for causing the antifertility and the neurotoxic effects.

Animals↗

PubMed1981

The effect of high doses of 6-chloro-6-deoxyglucose on the rat.

Male rats given 6-chloro-6-deoxyglucose (240 mg/kg/day for 28 days) developed spermatocoeles in their ductuli efferentes or caput epididymides. They had a lower serum triglyceride content than controls (0.87 +/- 0.19 vs 1.84 +/- 0.19 mM, Mean +/- SEM; n = 6) and gained less weight (2.55 +/- 0.37 vs 4.1 +/- 0.96 g/day, Mean +/- SEM; n = 6). There was no effect on female rats which received the same treatment. Spermatocoeles could also be produced by a single dose of 6-chloro-6-deoxyglucose, the threshold dose was between 180 and 240 mg/kg. Glucose oxidation by liver, brain, kidney and diaphragm from rats given 6-chloro-6-deoxyglucose (240 mg/kg/day for 14 days) was the same as in controls but was decreased in seminiferous tubules (0.32 +/- 0.06 vs 0.74 +/- 0.02 mumol [U-14C]glucose oxidised to 14CO2/g fresh wt/h, Mean +/- SEM; n = 3). The activity of glyceraldehyde-3-phosphate dehydrogenase [E.C. 1.2.1.12] in liver, brain, testis or muscle from rats given 6-chloro-6-deoxyglucose (24 mg/kg/day for 14 days) showed little change although its activity in spermatozoa was dramatically decreased.

Animals↗

PubMed1981

Effects of 6-chloro-6-deoxysugars on glucose oxidation in rat spermatozoa.

6-Chloro-6-deoxyfructose or 6-chloro-6-deoxyglucitol (greater than 90 mumol/kg/day), 6-chloro-6-deoxyglucose or 6-chloro-6-deoxymannose (greater than 120 mumol/kg/day) and 6-chloro-6-deoxygalactose (greater than 300 mumol/kg/day) all had an antifertility action in the male rat when given by mouth. Spermatozoa from the infertile rats were unable to oxidize glucose. This effect was always produced by a lower dose than the antifertility effect and the threshold dose for the 2 effects varied in a parallel fashion between the different 6-chloro-6-deoxysugars. Glucose oxidation appeared to be inhibited at the triose phosphate isomerase or glyceraldehyde 3-phosphate dehydrogenase reaction. These effects of 6-chloro-6-deoxysugars are similar to those of alpha-chlorohydrin. The activities of the glycolytic enzymes were greatly in excess of the observed flux through the pathway and high concentrations of glucose 6-phosphate and triose phosphate accumulated in rat spermatozoa incubated with 2 mM-D-glucose.

Animals↗

PubMed1981

The effect of 6-chloro-6-deoxysugars on adenine nucleotide concentrations in and motility of rat spermatozoa.

Spermatozoa from rats treated with 6-chloro-6-deoxyglucose (120 mumol/kg/day) were as motile as those from controls soon after dilution but rapidly became immotile when 3 mM-D-glucose was the only substrate present. Control spermatozoa were more motile when 2 mM-pyruvate + 4 mM-DL-lactate rather than 2 mM-D-glucose was present and the former substrate allowed spermatozoa from treated rats to remain motile. The concentrations of the adenine nucleotides declined rapidly during 40 min incubations when th substrate was 2 mM-D-glucose. The ATP/ADP ratio and the energy charge (ATP + 1/2 ADP/ATP + ADP + AMP) were maintained in spermatozoa from control rats but not in those from treated rats. When the substrate was 2 mM-pyruvate + 4 mM-DL-lactate adenine nucleotides were not lost as rapidly and there was less difference between spermatozoa from control and treated rats.

Adenine Nucleotides↗

PubMed1981

The role of oxidative phosphorylation in the generation of ATP in human spermatozoa.

Washed human spermatozoa had an endogenous oxygen uptake of 2.14 +/- 0.17 nmol O2/10(8) spermatozoa/min (mean +/- s.e..m., n = 35) which was stimulated by succinate (Vmax = 9.64 +/- 0.44 nmol O2/10(8) spermatozoa/min) but not by other substrates. The ATP concentration in freshly washed spermatozoa was 12.18 +/- 0.54 (s.e.m.) nmol/10(8) spermatozoa (n = 26) and was maintained for 2 h in the presence of 2 mM-D-glucose but fell to 9.56 +/- 0.73 (s.e.m.) nmol/10(8) spermatozoa (n = 13) in its absence. The presence of 2 microM-antimycin A, 2 microM-rotenone, 0.4 microM-carbonyl cyanide m-chlorophenyl hydrazone or 8 microM-oligomycin caused the ATP concentration to fall to less than 2 nmol/10(8) spermatozoa but their effect was partly alleviated by 2 mM-glucose. Sodium malonate (5 mM) prevented the stimulation of respiration by succinate but had no effect on the ATP concentration of the spermatozoa or their ability to produce 14CO2 from [U-14C]glucose. The least active of the tricarboxylic acid cycle enzymes was 2-oxoglutarate dehydrogenase (EC 1.2.4.2) (3.1 +/- 0.6 (s.e.m.) nmol substrate transformed/10(8) spermatozoa/h (n = 4). Cytochrome c oxidase (EC 1.9.3.1) was much less active than in rat spermatozoa (22.3 +/- 6.0 (s.e.m., n = 4) and 615 +/- 87 (n = 4) nmol transformed/10(8) spermatozoa/min). It is concluded that human spermatozoa can obtain ATP by the respiration of endogenous substrate but the substrates and metabolic pathways involved remain obscure.

Adenosine Triphosphate↗

PubMed1981

The neurotoxicity and antifertility properties of 6-chloro-6-deoxyglucose in the mouse.

6-chloro-6-deoxyglucose (24 mg/kg/day) produces infertility in male rats and marmosets. In the present study very high doses (480 mg/kg/day) were found not to impair the fertility of male mice. However, such doses induced the formation of vacuolated lesions in specific regions of central nervous system grey matter causing varying degrees of physical disability. A similar response has been obtained in marmosets. Astrocytes appear to be primarily affected with later involvement of nerve cells. The sites of lesions correspond closely to regions of high glucose utilization.

Animals↗

PubMed1980

The control of male fertility by 6-chloro-6-deoxysugars.

1. 6-Chloro-6-deoxyglucose (24 mg/kg/day) produces a rapid and reversible antifertility effect in the male rat but high doses (greater than 200 mg/kg/day) produce spermatocoeles. In the mouse repeated high doses (480 mg/kg/day) cause paralysis of the hind limbs but there is no effect on fertility. 2. Spermatozoa from rats treated with 6-chloro-6-deoxyglucose (24 mg/kg/day) are unable to metabolize glucose and rapidly become exhausted when glucose is the only substrate supplied. Glycolysis is inhibited at the glyceraldehyde 3-phosphate dehydrogenase reaction. These spermatozoa can obtain energy by oxidizing pyruvate or lactate. 3. The side effects prevent the use of 6-chloro-6-deoxyglucose by humans but a fuller understanding of their mode of action may lead to the development of compounds with a more selective attack on spermatozoa.

Animals↗