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Biomedical subjects

T M Mayhew

Publications and source records attributed to T M Mayhew.

At least 19 recordsLinked to original sources
PubMed1992

Structural and enzymatic studies on the plasma membrane domains and sodium pump enzymes of absorptive epithelial cells in the avian lower intestine.

The coprodaeum of the domestic hen maintained on a low-NaCl diet adapts by enhanced sodium transport. This study examines the adaptive response at the single cell and whole organ levels. Surface areas of apical (microvillous) and basolateral plasma membranes of columnar absorptive epithelial cells were estimated by use of ultrastructural stereology. The activities of succinic dehydrogenase (a mitochondrial enzyme) and ouabain-sensitive, potassium-dependent paranitrophenyl phosphatase (a sodium pump enzyme) were determined in tissue homogenates. Sodium, potassium-ATPase (pump enzyme) activity in cell membranes was localized by ultrastructural cytochemistry. Apical and basolateral membranes responded differently. In high-NaCl hens, the membrane signature of the average cell was 32 microns 2 (apical), 932 microns 2 (lateral) and 17 microns 2 (basal). Cells from low-NaCl hens had more apical membrane (49 microns 2 per cell) but essentially the same area of basolateral membrane. However, total surfaces per organ were greater for all membranes. Sodium pump enzymes were localized in basolateral membranes. Enzyme activities per unit mitochondrial volume and per unit basolateral membrane surface were higher in low-NaCl birds. These findings are discussed in the context of known mechanisms of transcellular sodium transport via apical ion channels and basolateral pumps.

4-Nitrophenylphosphatase↗
PubMed1992

A review of recent advances in stereology for quantifying neural structure.

The science of stereology has undergone a revolution over the past decade with the introduction of design-based (assumption- or model-free) methods which are highly efficient and generally unbiased. No other morphometric approach currently offers these twin benefits. Stereology is ideal for extrapolating 3-D structural quantities (real volumes, surface areas, lengths and numbers) from simple counts made on 2-D slice images. The images may take various forms (e.g. physical or optical sections, MRI slices, CT scans) but they must be sampled so as to be random in orientation and/or position if valid estimates are to be made. All the recent developments in stereology are applicable to problems in neuromorphometry. This review provides an account of major developments and the state of the art, emphasizes the importance of properly randomized sampling and identifies some applications to neural structure at different levels of organization. These include the counting and sizing of synapses, neurites, cells and whole brains.

Animals↗
PubMed1992

Responses of enterocyte microvilli in experimental diabetes to insulin and an aldose reductase inhibitor (ponalrestat).

The small intestine of 12-week-old streptozotocin-diabetic rats was examined by light and transmission electron microscopy in order to study the effects of alternative treatments on microvillous morphology. Four groups were examined: untreated diabetic rats, insulin-treated diabetics and rats treated with an aldose reductase inhibitor (ponalrestat) given with and without insulin. Numbers and dimensions of microvilli at the apex of columnar absorptive epithelial cells (enterocytes) were estimated using stereological principles. Values were obtained for the organ as a whole as well as for different sites along its length. In the untreated diabetic intestine, the mean (standard error of mean) number of microvilli was 4.5 (0.8) x 10(12) with a total surface area of 1.9 (0.50) m2. On average, the microvilli were 1.1 (0.08) microns long, 104 (3.8) nm in diameter and packed on the villous surface at a density of 3400 (50) per 100 microns 2. Their length at least varied with intestinal location. Significant effects of insulin therapy were detected. In contrast, the study failed to find any significant effect of aldose reductase inhibition on any variable except microvillous packing density.

Aldehyde Reductase↗
PubMed1992

Quantitative analysis of factors contributing to expansion of microvillous surface area in the coprodaeum of hens transferred to a low NaCl diet.

A stereological study of the lower intestine (coprodaeum) of the domestic hen was undertaken using combined light and electron microscopy. Numbers of columnar absorptive epithelial cells and the dimensions and numbers of microvilli were estimated. The aim was to identify the main factors contributing to an increase in microvillous surface area following transfer of hens from a high to a low NaCl diet on which they were kept for at least 3 wk. The principal contributor to observed changes between organs was cell number. Birds adapted to the low NaCl diet had 57% more cells than high NaCl controls. The average cell had a larger microvillous surface (55%) and this could be explained by the presence of longer (34%) and more densely packed (38%) microvilli. The total number of microvilli per coprodaeum doubled (from 35 x 10(9) on the high NaCl to 71 x 10(9) on the low NaCl diet). The increase in cell number accounted for 67% of the change in surface area and 78% of the change in number of microvilli per organ. These findings emphasise that, when assessing the form and function of a whole organ, it is important to monitor cell populations as well as single cells. This is especially true when studying renewing and expanding (rather than static) populations.

Animals↗
PubMed1992

A fractionator study of the effects of undernutrition during early life on rat Purkinje cell numbers (with a caveat on the use of nucleoli as counting units).

The effects of varying periods of general protein-calorie undernutrition during early life on Purkinje cells in the cerebellum of the rat were examined. In Experiment 1, animals were undernourished from d 18 of gestation until either d 30 or d 60 of postnatal age, followed in some cases by a period of nutritional rehabilitation. In Experiment 2, rats were undernourished from conception until d 30 postnatally, followed by a period of rehabilitation until 60 d of age. The 'fractionator' method was employed to estimate the total number of cerebellar Purkinje cell nucleoli. If each cell has one nucleolus, this number is equal to the number of Purkinje cells. In Experiment 1, the estimated mean number of Purkinje cell nucleoli varied between 188,000 and 273,000 for the groups of animals studied. However, the interanimal variation was large and 2-way analysis of variance tests failed to reveal any statistically significant age, nutrition or interaction effects. In Experiment 2, rats killed immediately after the period of undernutrition possessed significantly fewer Purkinje cell nucleoli than age-matched controls. This did not represent a real decrease in cell number. Instead, the result could be explained by a change of Purkinje cell morphology in undernourished rats. The change included nucleolar regression. Nutritionally rehabilitated animals had similar numbers of nucleoli to control rats. It is concluded that the levels and periods of undernutrition used in these experiments did not affect significantly the total numbers of cerebellar Purkinje neurons.

Animals↗
PubMed1991

Accurate prediction of Purkinje cell number from cerebellar weight can be achieved with the fractionator.

Purkinje cell nucleoli are used as counting units in order to obtain unbiased (fractionator) estimates of the number, N, of Purkinje neurons in adult mammalian cerebella of known weight, W. Regression analysis is then employed to establish the nature of the relationship between logN and logW. The linear regression equation defines an allometric relation that is employed to predict number in cerebella of known weight from other mammals. Predicted numbers are tested against empirical estimates. For 19 cerebella ranging in weight from 0.2 g (rat) to 113 g (human), the allometric relation between Purkinje cell number and organ weight was determined. By using this relation, the mean complement in three rabbit cerebella (average weight, 0.87 g) is predicted to be 0.63 million. This figure is confirmed by fractionator estimates made on the same three brains. The cat cerebellum should contain about 1.5-2.0 million Purkinje cells. An estimate of 1.2-1.3 million cells is to be found in the literature. Including rabbit cerebella in a refined equation yields the following relation: N = 686000W(0.695). With this refined equation, further predictions are made about the numbers likely to be found in the cerebella of the dog, goat, pig, ox, and horse. The numbers predicted for these animals must await experimental verification, but they are entirely consistent with previous suggestions that neuronal packing densities decrease with increasing brain size.

Algorithms↗
PubMed1991

Inhibitor action on placental calcium transport.

Human term placental lobules were dually perfused with Krebs Ringer solution at 37 degrees C under open circuit conditions. Provided that perfusate Ca2+ concentrations were between 2.33 and 2.55 mM, there was a steady release of Ca2+ into the fetal circulation and uptake of Ca2+ from the maternal circulation. There was no significant calcium (Ca) protein binding in the perfusates. Addition of dinitrophenol altered the release of Ca2+ to an uptake on the fetal circuit and enhanced Ca2+ uptake on the maternal circuit. It also produced a release of potassium (K)+ and an uptake of Na+ on both sides of the placenta. Ouabain had no significant effect on Ca movements although it produced a marked release of K+ into the fetal perfusate. The effect of cooling on the fetal circuit was similar to that of dinitrophenol (DNP), although it did not produce significant changes in either Ca2+ or K+ movements on the maternal side of the lobule. Both DNP and cooling reduced the Ca concentration ratio between fetal and maternal outflows to unity. Replacement of Na+ by choline Ringer had only transient effect on the extraction of 45Ca from fetal perfusate. These observations indicate that a Ca2+/Na (sodium)+ exchanger does not make a major contribution to the transplacental movement of Ca2+ from mother to fetus and that this process is more probably associated with membrane-bound ATPases.

Adenosine Triphosphatases↗
PubMed1991

Parathyroid hormone-(1-34) peptide activates cyclic adenosine 3',5'-monophosphate in the human placenta.

Dually perfused human term placental lobules were exposed to forskolin, bovine parathyroid hormone (bPTH(1-34)) and human parathyroid hormone related-peptides, hPTHrP(1-34), hPTHrP(67-86)NH2 or PTHrP(107-138) for 15 min in the presence of 3-iso-butyl-1-methyl-xanthine (IBMX); control lobules were exposed to IBMX alone. Homogenates of these tissues were then assayed for cyclic adenosine 3',5'-monophosphate (cyclic AMP) and results normalized per mg of protein. Exposure to forskolin or bPTH(1-34) on both sides, and exposure to bPTH(1-34) at a concentration of 30 nM on the maternal side of the placenta or 120 nM on the fetal side of the placenta, significantly enhanced tissue cyclic AMP production compared with tissue exposed to IBMX alone. Exposure to hPTHrP(1-34), hPTHrP(67-86)NH2 and hPTHrP(107-138) at a concentration of 30 nM on both sides of the placenta had no significant effect upon tissue cyclic AMP production.

Cyclic AMP↗
PubMed1991

Avian lower intestine adapts to dietary salt (NaCl) depletion by increasing transepithelial sodium transport and microvillous membrane surface area.

A tissue sampling scheme for tandem assessments of whole-organ physiology and ultrastructure was applied to the lower intestine (coprodaeum) of White Plymouth Rock hens on low- and high-NaCl diets. The objective was to correlate net amiloride-sensitive Na transport determined using the Ussing chamber with the plasma membrane surface areas due to microvilli at the epithelial cell apex. Hens kept on the low-NaCl diet for 3-4 weeks displayed a substantial increase in short-circuit current and in total microvillous membrane surface area. The latter rose from a group mean +/- S.E.M. of about 90 +/- 9.7 cm2 to one of 200 +/- 38 cm2 per organ. An increase in epithelial cell membrane contributed to, but did not fully explain, the increase in microvillous area. No differences in mean cell height or mean cell volume were found but the average cell in the low-NaCl birds was better developed in possessing a greater surface area of microvilli. On the high-NaCl diet, the epithelium was 33 +/- 2.7 microns tall and contained about 270 +/- 15 million cells. Each cell had a volume, on average, of 540 +/- 59 microns 3 and a microvillous surface of 32 +/- 2.6 microns 2. After NaCl depletion, there were 420 +/- 75 million cells and the average microvillous surface was 49 +/- 5.3 microns 2 per cell. The morphological adaptations alone do not explain the increased net Na transport found on the low-NaCl diet. Of cardinal importance is greater density of open Na channels in apical cell membranes.

Adaptation, Physiological↗
PubMed1991

Magnetic resonance imaging (MRI) and model-free estimates of brain volume determined using the Cavalieri principle.

A complete set of parallel (coronal) slices through a fixed human forebrain was generated by magnetic resonance imaging (MRI) and the Cavalieri principle, combined with point counting, was used to estimate brain volume. Alternative sampling schemes for estimating volume were then assessed by taking systematic and simple random selections of slices. Later, the brain was weighed and its fixed volume determined by fluid displacement. For the complete set of n = 28 MRI slices, the volume (1025 ml) was estimated with a coefficient of error (CE) of less than 1%. Decreasing the number of slices by systematic sampling increased the CE but this was still only 5% when just 5-6 slices were analysed. Estimated volumes varied from 947 ml to 1098 ml. Simple random sampling was less efficient (estimated volumes for 5-6 slices were 644-1187 ml). The forebrain actually weighed 1090 g and displaced 1060 ml of fluid. A set of 14 other brains was physically sliced in order to assess sampling errors in the context of observed brain-to-brain variation. It was found that 5-6 slices per brain is enough to yield efficient estimates of mean brain volume. The findings demonstrate the practicability of using MRI to estimate brain volumes unbiasedly and efficiently. The methods have great potential for noninvasive, longitudinal studies on in vivo brains and other organs.

Brain↗
PubMed1991

Scaling placental oxygen diffusion to birthweight: studies on placentae from low- and high-altitude pregnancies.

Stereological analysis of tissue sections has been used to estimate partial and total conductances of the human placental oxygen diffusion pathway. Analyses were undertaken for neonates and term placentae from populations living at low (400 m) and high (3600 m) altitude in Bolivia. Relationships between conductances and birthweight (and between trophoblast volume and birthweight) were explored by linear regression analysis and allometry. At both altitudes, the three partial conductances on the maternal side of the oxygen pathway scaled to birthweight in a similar fashion. This similarity did not extend to conductances on the fetal side. Beyond a limiting weight of roughly 3 kg, the highland fetus was disadvantaged in terms of its placental oxygen diffusive conductances and trophoblast volume. The findings support the notion that the structural adaptations seen at high altitude are more successful on the maternal rather than the fetal side of the placenta.

Adaptation, Physiological↗
PubMed1990

Striated brush border of intestinal absorptive epithelial cells: stereological studies on microvillous morphology in different adaptive states.

Different regions of small bowel were examined in five groups of rats in three separate experiments. The effects on mucosal morphology of position along the bowel, induced hypoproliferation (due to fasting), and induced hyperproliferation (due to streptozotocin diabetes) were investigated. Intestines fixed by in situ perfusion with buffered glutaraldehyde were sampled by strictly randomised procedures. Pieces of tissue from segments of roughly equal length were processed for electron microscopy and embedded in resin. Complete transverse sections were cut for light microscopy and estimates of villous surface areas were obtained by stereological methods devised for the purpose. Ultrathin sections from random sectors of the same tissue blocks were sampled systematically to obtain micrographs of the villous surface. These were analysed for quantitative information about microvilli (length, diameter, surface area, and number). Structural quantities from individual segments were pooled to provide values for the entire small bowel. Significant regional differences in villous and microvillous dimensions were found in all groups. The numbers of microvilli per bowel were remarkably constant in all control groups. Other variables were estimated reproducibly in rats of the same sex, strain, and average body weight. Effective absorptive surfaces did not show a linear gradient but tended to peak in middle segments. Neither fasting nor induced diabetes altered the mean length, diameter, or packing density of microvilli. However, surfaces due to villi and microvilli altered commensurately during fasting and induced diabetes. Therefore cell number seems to be the key quantity for determining villous and microvillous surface areas. The findings are discussed in the context of kinetic, biochemical, and physiological changes found in different adaptive states.

Adaptation, Physiological↗
PubMed1990

A sampling scheme intended for tandem measurements of sodium transport and microvillous surface area in the coprodaeal epithelium of hens on high- and low-salt diets.

A tissue sampling protocol for combined morphometric and physiological studies on the mucosa of the avian coprodaeum is presented. The morphometric goal is to estimate the surface area due to microvilli at the epithelial cell apex and the proposed scheme is illustrated using material from three White Plymouth Rock hens. The scheme is designed to satisfy sampling requirements for the unbiased estimation of surface areas by vertical sectioning coupled with cycloid test lines and it incorporates a number of useful internal checks. It relies on multi-level sampling with four levels of stereological estimation. At Level I, macroscopic estimates of coprodaeal volume are obtained. Light microscopy is employed at Level II to calculate epithelial volume density. Levels III and IV require low and high power electron microscopy to estimate the surface density of the epithelial apical border and the amplification factor due to microvilli. Worked examples of the calculation steps are provided.

Animals↗
PubMed1990

Fractionator studies on Purkinje cells in the human cerebellum: numbers in right and left halves of male and female brains.

Direct estimates of the numbers of Purkinje cell nucleoli in 22 aged human cerebella of known weight were obtained from paraffin-embedded sections using the fractionator. The nucleoli were counted in single sections. The estimates are unbiased by fixation, section thickness or sampling errors and are free from any assumptions about shape, size or spatial orientation. Left and right sides of the cerebellum were analysed in ten subjects whilst sex differences were examined in eleven subjects. No significant lateral differences in fixed weight or nucleolar number were observed. The group means (coefficients of variation) per side were 45 g (18%) and 7.8 millions (26%) respectively. The average complement of nucleoli in 22 cerebella amounted to 15.6 millions (22%). Estimated numbers did not show any significant correlation with cerebellar weights and, in a subsample of these brains (five male and six female), apparent sex differences were not significant.

Aged↗
PubMed1990

Comparative morphometry of the mammalian brain: estimates of cerebral volumes and cortical surface areas obtained from macroscopic slices.

Stereological methods for obtaining unbiased estimates of brain volumes and surface areas are illustrated on fixed brains (cerebrum only) from mammals varying in body weight between 2 kg (cat) and 550 kg (ox). Brain sampling was designed so that Cavalieri estimates of volumes (derived via point counting) could be combined with vertical sectioning estimates of surface areas (via intersection counting). Total volumes, cortical volumes and cortical surface areas were calculated. Each cerebral hemisphere was cut into 3 slabs. Each slab was cut further into macroscopic, parallel vertical slices having a uniform random start position. The term vertical as used here signifies orthogonal to the medial aspect of the hemisphere. The direction of vertical slices also had a random start but was varied systematically across the slabs. Test lattices bearing test points and cycloid test lines were superimposed on vertical slices. Fixed volumes and surfaces were corrected for shrinkage effects. A worked example of the calculation sequence is provided. The experimental design was flexible. Brains of different sizes could be analysed by simply altering the distance between slices and the size of the test lattice. Analyses took 30-45 minutes per hemisphere. Whilst volumes and surfaces increased with body weight, specific values declined. Thus, specific surfaces for the cortex fell from 25 cm2/kg (cat) to 2 cm2/kg or less (pig and ox).

Animals↗
PubMed1989

On the number of Purkinje cells in the human cerebellum: unbiased estimates obtained by using the "fractionator".

Stereological estimates of the numbers of Purkinje cell nucleoli in human cerebellar cortex have been obtained from systematic random samples of tissue by using the fractionator. The estimates are unbiased by fixation, section thickness, or sampling errors and are independent of any assumptions about cell shape, size, or spatial orientation. Twelve brains from aged subjects of both sexes were examined. The average complement of nucleoli in four female brains (age range 71-93 years) amounted to 14.8 millions (with an observed coefficient of variation between subjects of 29%). For three male brains (76-91 years), the corresponding estimates were 15.7 millions (10%). No significant sex differences were found for these small samples. Five brains of unknown sex and age yielded values of 15.8 millions (18%). For the twelve brains examined, the total number of Purkinje cell nucleoli per cerebellum was found to be 15.4 millions (19%). Estimated numbers showed a significant positive correlation with cerebellar weights. The number of nucleoli in an individual cerebellum was obtained with high precision in as short a time as 4 hours.

Aged↗