NADH-diaphorase Staining Research Articles

Human sperm nadh and nadph diaphorase cytochemistry: correlation with sperm motility

Objectives. We have examined the correlation between the retention of residual sperm cytoplasm and sperm motility in semen from men presenting for infertility evaluation. Methods. Semen samples (n = 12) were obtained from nonazoospermic men presenting for infertility evaluation at our institution. Samples were fractionated into high-, intermediate-, and low-density subpopulations by Percoll gradients in order to examine the correlation between the retention of residual sperm cytoplasm and sperm motility. Residual sperm cytoplasm retention was detected by cytochemical staining of sperm for nicotinamide adenine dinucleotide (NADH)- or nicotinamide adenine dinucleotide phosphate (NADPH)-dependent diaphorase activity. Results. The different sperm subpopulations (low, intermediate, and high density) had significantly different percentages of sperm with droplet retention (analysis of variance, P <0.05). Using either NADH or NADPH diaphorase staining as a marker of the cytoplasmic space, a significant negative correlation was observed between the percentage of sperm with residual cytoplasmic droplets and the percentage of motile sperm ( r = −0.58 and −0.61, respectively, P <0.05). Conclusions. Assessment of residual sperm cytoplasm retention is a simple diagnostic test. Although this test is of unproven value in the management of infertile men, this and other studies suggest that it may provide useful data on sperm function.

Quantitative study and architecture of nerves and ganglia of the rat heart.

The cardiac plexus of the Wistar rat was investigated in whole-mount preparations of the atria by NADH-diaphorase staining and by the acetylcholinesterase (AChE) histochemical technique. The plexus lies over the muscular layer of the atria, dorsal to the muscle itself, in the connective tissue of the subepicardium. The plexus contains on average 975 +/- 150 neurons, occurring singly or gathered in packed ganglia. The ganglia are found regularly in certain situations, namely, cranially to the pulmonary veins (44% of total); cranially and to the left of superior vena cava (10%); in the interatrial groove (21%); to the left of the left pulmonary vein (11%); caudally to the pulmonary veins (12%) and in the wall of the coronary sinus (1%). Ganglia are never found on the auricular appendages. For the histochemical demonstration of AChE, the 'direct coloring' copper ferrocyanide method was used. Extrinsic nerves enter the atria along the superior vena cava, along the pulmonary veins and along the coronary sinus, forming ganglion-containing plexuses. From specific sites of these plexuses, nerves proceed to the ganglia located to the left of the superior vena cava, to the ventral and dorsal ventricular wall and to the wall of the right atrium, where they form a delicate plexus accompanying the muscle fibers. Most of the neurons of the plexuses displayed AChE activity in the cytoplasm though they presented different reaction intensities. The distribution of cardiac nerves from groups of neurons located at discrete sites may indicate that postganglionic nerves selectively project to and thus control specific cardiac regions and/or functions.

Progressive reorganization of the myenteric plexus during one year following reanastomosis of the ileum of the guinea pig

The enteric nervous system appears to play a pivotal role in the functional recovery of the gastrointestinal tract after partial resection and reanastomosis, but the structural changes following surgery are not fully understood. The present study was designed to clarify the processes of myenteric plexus regeneration up to one year after transection and reanastomosis of the ileum of the guinea pig. The following techniques were used: nicotinamide adenine dinucleotide (NADH) diaphorase histochemistry, immunostaining of neuron-specific enolase (NSE) in whole-mount preparations, and transmission electron microscopy. Two months after transection and reanastomosis, myenteric ganglion cells with NADH diaphorase reactions were scarce in the center of the lesion, and were less numerous in adjacent areas (3 mm in width) than in the control ileum. In the areas adjacent to the lesion, a few large extraganglionic neurons that did not completely compensate for the loss of ganglion neurons were observed. The remaining ileum showed no changes in NADH diaphorase staining pattern at this stage. Two to 12 months after transection and reanastomosis, ectopic large neurons gradually increased in number not only in the areas adjacent to the lesion but also in part of the remaining ileum, up to 10 cm from the lesion. Concomitantly, large ganglion neurons decreased in number in these areas. In other ileal regions (more than 10 cm distant from the site of transection), no obvious changes in NADH diaphorase staining were noted throughout the observation period. The outgrowth of NSE-containing nerve fibers from the severed stumps was seen two weeks after transection. Six weeks later, numerous bundles of fine nerve fibers with NSE were shown to interconnect the oral and anal cut ends of the myenteric plexus, but they exhibited no subsequent alterations. Transmission electron microscopy revealed that regenerating nerve fiber bundles appeared initially among irregularly arranged smooth muscle cells eight weeks after the operation, as expected from light-microscopic observations. These findings suggest that myenteric ganglion cell bodies, unlike myenteric nerve fibers, require a longer term of reconstruction than previously believed after transection and reanastomosis of the ileum of the guinea pig.

Effects of surface electrostimulation on the structure and metabolic properties in monkey skeletal muscle

Adaptative changes in skeletal muscle following surface electrical stimulation (SES) were investigated in rhesus monkeys. SES was performed on the triceps brachialis muscle (TB) according to an intermittent pattern. The procedure was carried out for 3 wk, using a current with a medium frequency of 60 Hz normally observed in fast motor axons. The histochemical assays performed on biopsies taken from proximal and distal parts of the TB muscle, before and after the SES program, showed that the distribution of fibers typed by ATPase was unaffected. On the other hand, SES led to an overall increase in the mean fiber cross-sectional area (FCSA); P < 0.01 (+13.7%, NS, in proximal portion, vs +31%, P < 0.01 in distal portion). This increase in size occurred in all fiber types. SES was shown to induce an overall increase in capillary to fiber ratio (C/F; +11.06%, NS, in proximal portion, vs +25.93%, P < 0.05 in distal portion). The number of capillaries surrounding fiber Type II (CAFII) was significantly increased by SES (P < 0.05): +3.21%, NS, in proximal portion, versus +21.47%, P < 0.05 in distal portion. Moreover, the number of capillaries surrounding fiber Type I (CAFI) was statistically unaffected by SES. These results suggest that a stimulation of capillary growth may occur following SES-training. Citrate synthase activity was significantly increased after SES. This enhancement in oxidative potential was shown to occur in all fiber types (NADH-diaphorase staining).(ABSTRACT TRUNCATED AT 250 WORDS)

Fibre types in human lumbar back muscles.

The distribution of histochemically identified muscle fibre types was studied in biopsy samples from the two main muscles in the lumbar region of the human erector spinae, the multifidus and the longissimus, in 16 healthy subjects (nine males and seven females, age 20-30 years). Muscle fibres were classified as types I, IIA, IIB or IIC on the basis of the pH lability of their myofibrillar ATPases. There were no differences between the multifidus and the longissimus muscles in the relative occurrence of type I (62 vs. 57%), type IIA (20 vs. 22%) or type IIB fibres (18 vs. 22%), or in the absolute size of fibres (range of mean least diameters 58-66 micron). The oxidative potential (NADH-diaphorase staining intensity) was high in type I and low in type II fibres, irrespective of subgroups, in both muscles. In the females, the type I fibres occupied a relatively larger area (70-75 vs. 54-58% for the males) although the relative number of type I fibres was the same in both sexes. This was due to smaller type II fibres in the females resulting in higher type I/type II area ratios (1.70-1.90 vs. 0.88-0.92 for males). This suggests a difference in functional capacity of lumbar back muscles between males and females. On the other hand, the similarity in histochemical fibre-type distribution between the multifidus and the longissimus muscles does not give support for a functional differentiation between these two anatomically different parts of the lumbar erector spinae in man.

Diverse fiber composition of swimming muscles in the blue crab, Callinectes sapidus

The gross morphology and histochemistry of basal swimming muscles of the fifth pereiopod of the blue crab, Callinectes sapidus, were investigated. Each of the major muscles has two to three discrete branches and contains fibers ranging in colour from white to deep pink. The coloration is correlated with mitochondrial density. Deep pink fibers show intense NADH diaphorase staining, indicative of high oxidative capacity, while white fibers show very little staining, indicative of low oxidative capacity. Myrofibrillar ATPase activities are inversely correlated with oxidative capacities. The deep pink region of the levator muscle (muscle F) shows higher phosphorylase (glycolytic) activity, higher polysaccharide content and shorter sarcomere length than the white region. Such differentiation within a single muscle suggests considerable functional versatility.

Fibre types in human abdominal muscles

Histochemical muscle fibre composition was studied in biopsied from the four different muscles of the abdominal wall (rectus abdominis, RA, obliquus externus, OE, obliquus internus, OI, and transversus abdominis, Tr) in 13 normal human subjects (9 females and 4 males, age 24-55 years) undergoing gall-bladder surgery. Muscle fibres were classified as Type I, IIA, IIB or IIC on the basis of their myofibrillar ATPases' pH lability. There were large inter-individual variations in fibre composition, whereas, in general, the differences between the different muscles were minor or non-existent. Mean fibre distribution ranges were 55-58% I, 15-23% 22A, 21-28% IIB, and 0-1% II C fibres. The least fibre diameters were similar for all types and muscles (range of means 50-54 micrometer) except for Tr in which the Type II fibres were smaller (mean 45 micrometer). There was a high correlation in the size of Type I vs. II fibres and Type IIA vs. IIB fibres in all layers. The oxidative potential (NADH-diaphorase staining intensity) appeared high in Type I fibres and low in Type II fibres, irrespective of subgroups. Thus, based on histochemical fibre composition, the different abdominal muscles appear to have a similar functional capacity. However, functional differences between individuals were indicated by the large inter-individual variation in muscle fibre distribution.