Andrew J. Lechner

Lung morphometry in guinea pigs acclimated to hypoxia during growth.

The effect of chronic hypoxic exposure on lung development has been assessed in growing guinea pigs (Cavia porcellus). Weanling males of initial W = 229 g were acclimated to a PO2 of 80 Torr for 2-14 weeks before sacrifice (range of W = 244-965 g). Growth was the same in hypoxic animals as in controls maintained at a PO2 of 133 Torr (range of W in controls = 89-1274 g). Lungs were fixed by tracheal instillation of glutaraldehyde and examined morphometrically with the electron microscope. Within 3 weeks of exposure, lung volume (VL) and alveolar surface area (Sa) were significantly increased by 32% and 27% respectively in the hypoxia acclimated animals compared to controls of similar W. However, these differences were progressively reduced with increasing time of exposure, and mean values of VL and Sa were not different between groups when W greater than 900 g. Chronic hypoxia accelerated lung development towards normal adult dimensions to a degree remarkably similar to that reported in cold acclimated guinea pigs. These findings are compatable with the theory of adaptive lung growth mediated by increased pulmonary blood flow, and suggest anatomical limitations to such growth related to an animals age.

The closest-individual method in the analysis of the distribution of capillaries ☆

Abstract An ecological survey method termed the closest-individual method was adapted for measuring the distribution of capillaries in tissue. Models were constructed in which dots represented transversely sectioned capillaries in known densities, and in ordered, random, or contagious (clustered) array. The distances from randomly selected points on a model to the nearest capillary were measured, and the frequency distributions of these distances were plotted. This distribution was compared to the mathematically predicted distribution of distances for each model. All values derived by the closest-individual method were statistically similar to predicted values, and were within approximately 5% of predictions. In addition, the kurtosis of each set of data was used to evaluate whether the data from the closest-individual method could correctly identify the nature of the array of capillaries in each model. This method is potentially of great use to studies of capillarity since it calculates median diffusion distance more accurately than estimates from capillary density alone; it calculates maximal diffusion distance, which cannot be estimated from capillary density alone; and by identifying the array of capillaries as either random, ordered, or contagious, it assesses the nature of interaction among capillaries. Other similar measurement methods are discussed and compared.

Increased capillary supply in skeletal muscle of guinea pigs acclimated to cold

The ATPase technique was used to visualize blood capillaries and to study fiber composition in 10-micrometer transverse sections of guinea pig gastrocnemius and soleus muscles. A control group of newborn, weanling, juvenile and adult male guinea pigs (GP) (BW = 89-1274 g) was studied in a 20-24 degrees C environment (22 degrees C GP) while 2-3 week old animals were exposed continuously to 5 degrees C for 2-18 weeks before sacrifice (5 degrees C GP) (BW = 239-1074 g). Body weight gain was not affected by cold exposure; however, the gastrocnemius and soleus muscles of the 5 degrees C GP grew at a slower rate than did the muscles of the 22 degrees C GP. The equations relating fiber cross sectional area (FCSA) and muscle weight (MW) were not different between the 22 degrees C GP and 5 degrees C GP for the soleus and gastrocnemius. Therefore, in both muscles at the same BW, FCSA was smaller in the 5 degrees C GP than in the 22 degrees C GP. In both of the two muscles of each group, capillary density (CD) decreased hyperbolically with increasing FCSA, while the capillary to fiber ratio (C/F) and the average number of capillaries around each fiber (CAF) increased linearly with increasing FCSA. The regression lines for CD, C/F and CAF versus FCSA for both muscles were parallel between groups, but at any FDSA, the CD, C/F and CAF were greater in the 5 degrees C GP than in the 22 degrees C GP. Percent fiber composition of the gastrocnemii of the 22 degrees C GP and 5 degrees C GP were not different; however, at the same FCSA each fiber type had a greater capillary supply in the 5 degrees C GP. The increased capillarity in the gastrocnemius and soleus muscles of the 5 degrees C GP suggests an improved capacity for oxygenation, a response which would correlate well with the increased oxygen utilization during prolonged cold exposure.

Increased capillarity in skeletal muscle of growing guinea pigs acclimated to cold and hypoxia.

Capillarity was evaluated on transverse sections of frozen gastrocnemius and soleus muscles of young, growing guinea pigs exposed to the combined stresses of cold (6 degrees C) (C) and hypoxia (ambient PO2 = 85 Torr) (H) for up to 16 weeks and these data were compared to those obtained in a control group of guinea pigs kept in Denver (22 degrees C, ambient PO2 = 133 Torr). Capillarity was assessed from measurements of capillary density and capillarity density to fiber density ratios. Mean (R) and maximal (R95) diffusion distances were measured by the closest individual method. The body growth rate of guinea pigs exposed to C + H was the same as that in the control condition. The gastrocnemius muscle grew at the same rate as in the control guinea pigs. Exposure to C + H produced a significant (P less than 0.001) increase in the capillary density and the C/F of the gastrocnemius, reducing the mean and the maximal diffusion distances. However, the soleus muscles of the guinea pigs in C + H did not grow at the same rate and relative to body size the soleus muscles of these guinea pigs in C + H were smaller due to their smaller fiber cross-section area; consequently, there was a relatively larger capillarity in these muscles. It is hypothesized that the increased muscle capillarity in animals exposed to C + H results from a marked lowering of the tissue PO2 which may result from a leftward shift of the Hb-O2 dissociation curve.

Evaluation of the concentric-circles method for estimating capillary-tissue diffusion distances

Abstract The concentric-circles method for estimating the complete distribution of distances from any point in the tissue of animals to the nearest capillary was evaluated. Models representing capillaries in transverse sections of muscle tissue were devised, using dots at various densities in ordered, random, or contagious (clustered) arrays. The distribution of distances from random points to the nearest capillary in each model was derived mathematically. These values were compared to values estimated by a method using sets of concentric circles placed over the models. In each set of circles, the smallest circle containing a capillary was recorded. The concentric-circles method generated estimates of median and maximal distances to the nearest capillary, and provided complete distributions of distances that were statistically similar to predictions, and deviated from predictions by an average of less than 10%.

The distribution of diffusion distances in the gastrocnemius muscle of various mammals during maturation.

Large changes in fiber size, capillary density and capillary/fiber architecture occur during maturation of mammalian skeletal muscle. To examine the effect of these changes on oxygen diffusion distance, the mean (R) and maximal (R95) distances from the capillary to the tissue were measured in the gastrocnemius of maturing guinea pigs, rats and dogs. Hyperbolic relationships between capillary density andR and between capillary density andR95 were found for the combined data from these three species. TheR andR95 increased with decreasing CD in growing guinea pigs and rats, but they remained constant in dogs. Statistical analyses showed that the capillaries in this muscle in all three species tended to be distributed in ordered arrays. These anatomical measurements are discussed in relation to their potential physiological impact on oxygen delivery to tissues.

Hematology in developing guinea pigs—I. normal maturation

Abstract 1. 1. Newborn to 24 week old male guinea pigs were studied at 22°C and an ambient Po2 of 133 torr. The following hematological variables were measured once on each animal: hematocrit, [Hb], RBC indices, WBC differentials, and the molar ratio of 2,3-diphosphoglycerate (2,3-DPG) tq [Hb]. 2. 2. [Hb] declined from 15.8 g/100ml at birth (BW = 90 g) to 12.6 g/100 ml when BW = 252 g, and then increased linearly with increasing BW. Red cell indices were nearly constant at all BWs: MCV = 81.7 μm3; MCH = 25.1 pg; and MCHC = 30.7%. In newborn guinea pigs, lymphocytes comprised the majority (73%) of WBCs, but this percentage fell to 55% in mature animals. The ratio of [2,3-DPG]/[Hb] rose from 1.40 M/M following birth to 1.95 M/M (BW = 154 g) and then declined to 0.92 M/M in mature animals. 3. 3. The relative maturity of the hematological profile in newborn guinea pigs contrasts with those in the laboratory mouse and laboratory rat.

Hematology and red cell morphology in guinea pigs acclimated to chronic hypoxia during growth

Abstract 1. 1. Weanling male guinea pigs were exposed for up to 14 weeks to an ambient P o 2 of 80 torr. Hematological measurements made on these animals were compared to data for normoxic controls of similar age and body weight. 2. 2. Within 2 weeks of hypoxic exposure, hematocrit was increased by 42% and [Hb] by 41% compared to controls. However, these changes were accompanied by only a 13% increase in the No. RBCs/mm 3 during the same period; mean red cell volume and mean cell hemoglobin content were each increased by 22% compared to controls. Normochromic macrocytes as well as numerous nucleated RBCs were evident in the peripheral smears of the hypoxia acclimated animals. 3. 3. Frequency analysis of RBC diameters showed a single peak at 6.3 μm for all controls. In guinea pigs acclimated to P O 2 = 80 torr, RBC diameters were initially bimodally distributed, with peaks at 6.3 and 7.6 μm, and later were broadly distributed with a mean of 7.0 μm. 4. 4. These findings support the hypothesis that, under severe and chronic erythropoietic stress, normochromic macrocytes appear in the peripheral blood following a skipped terminal mitotic division, during which cytoplasmic volume would normally be reduced.

Hematology in developing guinea pigs—II chronic exposure to 5°C

1. 1. Weanling male guinea pigs were exposed for up to 18 weeks in environmental chambers to 5°C and an ambient Po2 of 133 torr before sacrifice (range of BW at 5°C = 239−1,074 g). Growth rates of animals raised at 5°C were not significantly different from animals maintained at 22°C. Hematological measurements were made on each animal, including hematocrit, [Hb], RBC indices, WBC differentials, and the molar ratio of 2,3-diphosphoglycerate (2,3-DPG) to Hb. 2. 2. Cold exposure results in significant (P < 0.001) increases of 6–12% in Hct and [Hb], due to greater No. RBCs, compared to controls of similar BW. Within 2 weeks of exposure to 5°C, the 2,3-DPG/Hb ratio rose transiently to 1.84 M/M, and then was constant at about 1.40 M/M in animals exposed to 5°C for longer times. When compared to values for adult controls of 0.92 M/M, such an increase in the 2,3-DPG/Hb ratio would increase the P50 of blood from cold acclimated guinea pigs by 5–6 torr. 3. 3. The significant increases over control values in circulating [Hb] and in 2,3-DPG would substantially improve tissue O2 delivery in cold acclimated guinea pigs, by facilitating the transport and unloading of O2.

The hematological response to hypoxia in growing guinea pigs is blunted during concomitant cold stress

Abstract 1. 1. Weanling male guinea pigs were exposed for up to 16 weeks to the combined stresses of cold (6 C) and hypoxia (ambient P o 2 = 85 Torr). Hematological data for these animals and a group of adults maintained in the same environment for 4 weeks were compared to similar data for age and weight-matched controls raised at 22°C and P o 2 = 133 Torr. 2. 2. During the initial 6 weeks of cold & hypoxia, hematocrit and [Hb] were elevated by 10–13°, above controls; these increases were accompanied by nonsignificant changes in #RBCs/mm 3 , so that mild macrocytosis was evident. 3. 3. From week 6 onward, the hematological profile changed more dramatically, so that by week 16. the Hct, [Hb], and #RBCs were 31, 35 and 22% greater than controls respectively. 4. 4. Throughout the 16 weeks, polychromasia was slight and nucleated red cells were rare in the peripheral smears. 5. 5. These data for the weanling guinea pigs as well as those for the adult group indicate that particularly during the initial stages of exposure, concomitant cold stress appeared to blunt the erythropoietic response when compared to that reported earlier for a similar group of animals raised under a similar level of hypoxia but at 22°C.