Histochemical study on the distribution of some enzyme activities in the vagal and facial lobes of the goldfish, Carassius auratus.

The histochemical localization of six enzymic activities (acetylcholinesterase, pseudocholinesterase, monoamine oxidase, lactate dehydrogenase, succinate dehydrogenase and glucose-6-phosphate dehydrogenase) has been studied in the vagal and facial lobes of the goldfish, Carassius auratus. These encephalic centers are hypertrophic in Cyprinidae, corresponding to the dominance of gustatory function. Acetylcholinesterase shows a complex laminar distribution in the vagal lobes and a peculiar cellular localization in vagal motor neurons. Monoamine oxidase activity is mainly evident in fibrous tracts coming to or leaving from the lobes. Among oxidative enzymes examined, lactate dehydrogenase and succinate dehydrogenase exhibit distribution patterns respectively similar to those observed for acetylcholinesterase and monoamine oxidase. Some features on enzymes distribution in the gustatory centers of Carassius are in agreement with the enzymatic patterns well known in higher vertebrates.     

Myonic makeup of the muscles innervated by the vagus nerve

Histochemical investigation on succinic dehydrogenase activity and morphometric studies have demonstrated certain differences in the dog sublingual group of muscles. The thyreohyoid and sternohyoid muscles innervated by spinal nerves possess three types of myons differing in succinic dehydrogenase activity and in the area of transversal section. The cricothyreoid muscle and the superior pharyngeal constrictor obtaining their motor innervation from the vagus nerve are composed of unitypical muscular fibres with nearly the same areas of transversal section and high enzymic activity. The differences noted should be explained by different sources of motor innervation.     

Succinate dehydrogenase localization in the mitochondria of the renal tubules of cold- and warm-blooded vertebrates

Using electron microscopic histochemical technique, studies have been made on the activity of succinic dehydrogenase in the kidneys of the cod Gadus morrhua and dog. It was shown that chelate granules indicating localization of the enzyme in the mitochondria of nephronal cells, concentrate mainly in two zones -- between the membranes and inside the cristae. This distribution of the enzyme implies the presence of two pools of succinic dehydrogenase in the mitochondria which are utilized at different stages of oxidative phosphorylation. Succinic dehydrogenase content of the cristae is lower in cod than in dog.     

Quantitative histoenzymatic analysis of the hypothalamic neurosecretory system in rat ontogeny

In experiments on 6 and 16 days old rats cytophotometric studies have been made of histochemical reactions for succinate, lactate and glutamate dehydrogenases, alpha-glycerophosphate dehydrogenase and glucose-6-phosphate dehydrogenase in the supraoptic nucleus, paraventricular hypothalamic nucleus and the posterior hypophysis. It was found that heterochronous development of the neurones in the supraoptic and paraventricular nuclei, as well as the development of their axons in the posterior hypothalamus depend on the rate of maturation of the enzymic systems in postnatal life. In consolidation of the unique structure of the hypothalamic neurosecretory system, the key role is played by afferent influences, succinic dehydrogenase being involved into their realization. The data obtained indicate the importance of heterochronous development of the enzymic activities in the formation of bifunctional properties of neurosecretory hypothalamic neurones and reveal the primary development of neurotransmittery function as compared to the excretory one.     

A Histochemical Study of Normal and Denervated Red and White Muscles of the Rat

The distribution and characterization of the fibers of normal and denervated red and white muscles of the albino rat are reported in this study. Histochemical procedures for succinic dehydrogenase, lipides, adenosinetriphosphatase, esterase, and glycogen were utilized to differentiate muscle fibers, and these methods facilitated the study of the distribution of fiber types within whole muscle. Muscle fibers of the granular type (dark or red fibers) can be clearly distinguished from those with clearer sarcoplasm (light or white fibers) by methods for demonstrating succinic dehydrogenase, lipides, and esterase. The method for adenosine-triphosphatase reveals differences only under the special conditions described in the text. Additional fiber types are described in the cat's diaphragm and in the extrinsic ocular muscles of the rat. Succinic dehydrogenase and adenosinetriphosphatase activities of the soleus and biceps femoris were studied 14 days after denervation of these muscles. The histochemical findings are discussed principally in the light of current biochemical knowledge of these enzymes.     

A histochemical study on the correlation between secretory activity and the TCA cycle in the gland cell

Summary The activity of succinic dehydrogenase and malic dehydrogenase was observed histochemically in the gland stomach of rats, and also the relationship between the secretory activity of the gastric gland cells and the process of the TCA cycle in the cells was studied.Histochemically, enzyme activity is plainly visible in the gastric parietal cells but in the gastric chief cells and mucous neck cells.The secretory activity of the cells was promoted by the administration of food, the sub-cutaneous injection of histamine, histidine, acetylcholine or eserin.The activity of succinic dehydrogenase appears to be constant regardless of secretory activity except in a few cases. The activity of malic dehydrogenase increases as secretory activity is promoted. It seems very unlikely that one step in the cycle (the transformation of malic acid into oxalacetic acid) would be accelerated while the other step (the transformation of succinic acid into fumaric acid) is not. This inconsistency of activity may be attributed to the histochemical reaction. Thus the increase of malic dehydrogenase activity is seen as an acceleration of the whole TCA cycle. It is our conclusion, therefore, that the source of energy within the cell, i.e. the TCA cycle, is a process which parallels secretory activity.     

Histochemical localization of enzyme activity during floral evocation in the shoot apical meristem ofSinapis alba

Summary Vegetative plants ofSinapis alba, a long-day species, were induced to flower by exposure to a single 20-hours day. Acid phosphatase, ribonuclease and succinic dehydrogenase activities were investigated by histochemical procedures at different times during floral evocation of the shoot apical meristem. There was an increase in reaction intensity for the three enzymes. Stimulation of acid phosphatase activity began at the 14th hours after the beginning of the long day; ribonuclease at the 18th hours, and succinic dehydrogenase at the 22nd hours. For the first two enzymes, activities returned to control values by 54 hours whereas succinic dehydrogenase activity was still increasing at 54 hours. Results are discussed in relation to other events which are known to occur in the meristem ofSinapis during the transition from the vegative to the reproductive condition.     

Histochemistry of the glycogen body of the turkey spinal cord

Summary Enzyme histochemical studies of the glycogen body of the turkey showed very little activity of suocinic dehydrogenase and cytochrome oxidase in the glycogen body cells, and marked activity of lactic dehydrogenase, NAD-diaphorase and the hexosemonophosphate shunt enzymes. Gradients of histochemical staining intensity for lactic and succinic dehydrogenase in the glycogen body and spinal cord were confirmed by biochemical assays of homogenates of these tissues. It was concluded that glycogen body metabolism is predominantly glycolytic. Alkaline phosphatase activity was weak; acid phosphatase activity was moderate. There was no acetyl cholinesterase or nonspecific cholinesterase activity in the glycogen body.This investigation was supported by U. S. Public Health Grant B 3250.Submitted in partial fulfillment of Masters' degree requirements.     

A histochemical study of cervical motor neurons and the posterior latissimus dorsi muscle in normal and dystropic chickens.

A chromatolysis study, 14 to 21 days following denervation, showed the spinal cord representation of the nerve to the posterior latissimus dorsi muscle to be in the ventrolateral cell column between cervical ganglia 14 and 15. to characterize cevical neruos nt undergoing chromatolysis, histochemical stuies were done the cords of additional nondenervated animals. Staining reactions for beta-hydrocybutyrate dehydrogenase, succinic dehydrogenase and cholinesterase did not reveal any quantitative differences between motor neurons in cervical segments 14 and 15 of normal and dystrophic birds. Motor neurons are positive for beta-hydroxybutyrate dehydrogenase and succinic dehydrogenase, but the surrounding neuropil is positive for the latter only. No pseudocholinesterase activity is found in the ventral horn cells, but true cholinesterase is present in most of the neurons...     

Histochemical studies on the olfactory glomeruli of squirrel monkey

Summary Detailed histochemical studies on the distribution of various oxidative and dephosphorylating groups of enzymes have been made in the olfactory glomeruli of the squirrel monkey. The olfactory glomeruli showed strongly positive activity for succinic dehydrogenase, lactic dehydrogenase, monoamine oxidase, alkaline phosphatase, adenosinetriphosphatase and simple esterase. They showed moderately positive activity for cytochrome oxidase, specific cholinesterase, 5'nucleotidase; mildly positive activity for acid phosphatase; and negligible activity for nonspecific cholinesterase and glucose-6-phosphatase. The glomeruli did not show the presence of any thiamine pyrophosphatase-positive Golgi apparatus. The blood vessels surrounding the glomeruli were strongly positive for the nonspecific cholinesterase test. The significance of these results are discussed briefly.     

Histochemical studies of several "K"-cell enzymes in guinea pig thyroid glands]

The thyroid gland of guinea pigs were studied morphologically. Histochemical methods were used for detection of lactate dehydrogenase, succinic dehydrogenase, cholinesterase, alkaline phosphatase and acid phosphatase. The distribution of "C"-cells in normal thyroid glands was proved to be uneven. In the center of the gland they were more numerous. For statistical investigations the method of silver impregnation of "C"-cells is more practicable, since they can not be obviously distinguished from acinar cells on the basis of glycerophosphate dehydrogenase only. The activity of cholinestarase in "C"-cells and in some other cells of folliculi epithelium is very high. A supposition is made that there exist two kinds of the follicular lining thyrocytes, having different histochemical properties and histogenesis as well.     

Comparative enzyme histochemical studies on the development of teeth in man and mouse

Summary On the basis of a material consisting of primordia of deciduous teeth from 17 human foetuses and tooth germs from 20 newborn mice and 4 mouse foetuses, the histochemical distribution of succinic dehydrogenase, LDH, BDH, glucose-6-phosphate-dehydrogenase, NADH₂ cytochrome C reductase, and NADPH₂ diaphorase was studied.The investigation comprised partly mapping of the above-mentioned oxidative enzymes in human tooth germs during the period from bud stage to bell stage with incipient hard-tissue formation, and partly a comparative enzyme histochemical investigation of the dental development in man and in the mouse.In both materials there was an increase of all the demonstrated enzyme systems in the ameloblasts in early amelogenesis. On the other hand, the ameloblasts of the two materials showed differences during the period of matrix production: In the human ameloblasts there was activity of succinic dehydrogenase basally as well as apically, while the non-human ameloblasts showed such activity only basally.The odontoblasts of both materials showed a definite reaction for glucose-6-phosphatedehydrogenase, LDH, and BDH, but slight activity of succinic dehydrogenase. This indicates the existence of the pentose shunt as the source of energy in dentinogenesis. On the basis of the enzyme reactions studied, neither material showed a subodontoblastic layer or any reaction in the hard tissues.Material supplied by the Association for the Aid of the Crippled Children, New York. Aided by grants from the Danish State Research Foundation and the Rask-Ørsted Foundation.     

Histochemical fibre types in the lateral muscle of fishes in fresh, brackish and salt water

The histochemical pattern of red, pink and white muscle of fish living in fresh, brackish, and salt water is reported. The muscle fibres were stained routinely during the year for lactate dehydrogenase (LDH), menadione α-glycerophosphate dehydrogenase (Mα—GPDH), succinic dehydrogenase (SDH), myosin adenosine triphosphatase (myosin ATPase), phosphorylase, lipids and glycogen. The pink and red muscles contain more glycogen and lipids and have a higher SDH activity, which is in accord with their aerobic metabolism and function in sustained swimming activity. The acid labile myosin ATPase activity characteristic of fast twitch fibres is present in the white fibres of most species, however in the white muscle of Gobius paganellus the enzyme activity is stable to both acid and alkali and, in addition, there is a scattered distribution of different fibre types in red and, especially, pink muscle. A study of seasonal variation patterns of myosin ATPase in white muscle of mugilidae over a period of two years has demonstrated, in late summer, the appearance of new small diameter fibres, with a high acid stable enzyme activity, that develop into the large diameter acid labile fibres.     

Electron transporting enzymes of rheumatoid connective tissue

The oxidative enzyme system of the synovial membrane and rheumatoid node from rheumatoid arthritic patients has been studied by histochemical and biochemical methods. As compared to the control synovial membranes, succinic dehydrogenase activity was substantially higher in the rheumatoid arthritic synovial membranes, and was still higher in the rheumatoid nodes, in which only the cells forming the palisade had succinic dehydrogenase activity. As compared to the controls, in response to menadione succinic dehydrogenase was significantly activated. The reducible ubiquinone content of the rheumatoid synovial membrane and the rheumatoid node was by several orders of magnitude higher than in the controls.     

Comparative microspectrophotometric study of the activity of oxidative enzymes in separate types of fibers of the skeletal muscle of rats in the normal state and in experimental ischemia]

Microspectrophotometrical investigation of the activity of oxidative enzymes (glutamatdehydrogenase, cytoplasmic and mitochondrial alpha-glycerol phosphate dehydrogenase, succinic acid dehydrogenase and glucose-6-phosphatedehydrogenase) in certain types of muscular fibres of m. soleus, gastronemius and quadriceps of rats during experimental stenosins of the abdominal part of the aorta has shown the existence of certain cyclic changes in the enzymic activity of different periods after graded constriction which may be associated with arising of adaptational reactions of metabolic systems in striated muscles in response to chronic hypoxia.     

Regional histochemical aspects of xanthine oxidase activity in ischemic and reperfused small intestine of the rat

The study describes regional changes of xanthine oxidase and succinate dehydrogenase activities as shown by the ischemic and reperfused small intestine of the rat. The results are obtained with enzyme histochemical methods, including densitometrical verifications, and are substantiated with biochemical enzyme determinations. The decrease of xanthine oxidase activity was best visible in the anoxic duodenum and jejunum, where the findings of histochemical enzyme determinations agreed with those achieved biochemically. The activities of succinate dehydrogenase as measured densitometrically may serve as a further control, considering also the typical intracellular distribution of the reaction products.     

A brachial glycogen body in the spinal cord of the domestic chicken.

When cervical segments 14 to 15 of the chicken spinal cord are cut transversely and studied by routine histological and histochemical methods, an onion-shaped region, filled with thread-like fibers, is seen to surround the ependymal cells of the central canal and to be bounded laterally by the neural elements of the spinal gray matter. This area is negative for succinic dehydrogenase, beta-hydroxybutyrate dehydrogenase and cholinesterase activity, but very strongly periodic acid-Schiff positive. Diastase controls show the positive material to be glycogen. Parasagittal sections through this cervical region and into the upper thoracic cord, show the glycogen-rich region to extend longitudinally throughout the region. Because of its location and histochemical characterization, which are similar to that of the ventral portion of the glycogen body, the term brachial glycogen body is proposed for this structure.     

Structural and metabolic changes in the limbic cortex of the rabbit in response to experimentation with the hypothalamus

By means of the light and electron microscopy, histochemical and cytophotometrical methods the anterior subarea of the cerebral limbic cortex has been studied in 30 experimental and 30 control rabbits. The experimental animals have been given 3, 15 and 30 sessions (1 h per day) of electric irritation (0.05 mA, 50 Hz, 1 msec) on the posterior hypothalamic field (PHF). Twelve rabbits from 30 control animals make an intact group and 18--a group with inactive electrodes inserted into the PHF. After 3 and especially after 15 sessions it has been revealed: in neurons--an acute swelling, edematous alterations, hyperchromatosis and shrinkage, changes in lactate dehydrogenase, succinic dehydrogenase, nicotinamide-adenine-dinucleotide-diaphorase and nicotinamide-adenine-dinucleotide-phosphate-diaphorase activities, in neuroglia--hypertrophy and weakly manifested hyperplasia. After 30 sessions synapses degenerated after the dark type are revealed. More intensive structural and metabolic changes are noted in the middle cytoarchitectonical complex. The changes of the enzymatic activity are considered as certain signs of weakening mitochondrial processes, connected with energy production, increasing glycolysis, decreasing level of the energetic provision of the cytoplasmic synthesis, arising under conditions of a disturbed transneuronal influence on the cortex by the hypothalamus.     

Demonstrability of some oxidative enzymes in early rodent embryos with and without fixation

Several oxidative enzymes [NADH-TR (reduced nicotinamide-adenine dinucleotide-tetrazolium reductase), NADPH-TR (reduced nicotinamide-adenine dinucleotide phosphate-tetrazolium reductase), SDH (succinic dehydrogenase) and LDH (lactate dehydrogenase)] were studied by histochemical means during early development of rat and mouse. All investigated enzymes could be easily demonstrated in zygote and also to some extent in somitic stages without any pretreatment. However, in cleavage and early postimplantation stages enzyme activity could be revealed only after the embryos were pretreated in some way. This pretreatment can be fixation with formalin or acetone, freezing and thawing, slight mechanical damage or very prolonged incubation time. The formazan granules as a sign of enzymatic activity were present in all stages of embryonic development and were more abundant in reactions for NADH-TR and LDH than in reactions for NADPH-TR and SDH. Our results suggest that the investigated enzymes are present in all embryonic cells during early development. It seems that the permeability of embryonic cells for histochemical media must be increased otherwise the histochemical reactions cannot be accomplished.     

The Histochemical Localization of Triphosphopyridine Nucleotide Diaphorase

A histochemical method is described for the localization of triphosphopyridine nucleotide diaphorase using a recently synthesized tetrazolium salt (Nitro-BT). By virtue of the favorable histochemical properties of this reagent, it has been possible to demonstrate that whereas DPN diaphorase is usually restricted to the mitochondria, the TPN diaphorase activity of corresponding cells was distributed throughout the cytoplasm in granules too fine to be considered mitochondria. Furthermore, although the diaphorase alone is responsible for the passage of electrons from TPNH to the tetrazole, it has been found that sites of activity of different TPN-linked dehydrogenases can be visualized in tissue sections, and characteristic loci for each enzyme may be observed. For example, whereas TPN diaphorase and isocitric dehydrogenase have an extensive distribution in the kidney cortex, 6-phosphogluconic dehydrogenase is limited to the cells of the macula densa.