Intrinsic neuromuscular defects in the neurogenic bladder: X. Value and limitations of neurohistochemistry

Histochemical changes in bladder innervation have been described in various types of clinical and experimental neurogenic voiding dysfunction. Recent studies in the cat have shown that both decentralization and postganglionic “denervation” lead to ultrastructural changes in muscular innervation of the bladder base. The present study was conducted to find out whether these changes can be detected histochemically. Decentralization of the bladder by unilateral sacral ventral rhizotomy, bilateral sacral ventral rhizotomy, or unilateral pelvic neurectomy was performed in 25, and postganglionic “denervation” by unilateral pelvic plexus neurectomy in 9, adult male cats. Specimens of the bladder body and bladder base were collected 2–4 and 8–10 weeks after decentralization, and 2–4 weeks and 10 weeks after “denervation.” The specimens were processed by histochemical methods to demonstrate cholinergic and adrenergic nerves. No recognizable differences from the norm were observed in cholinergic muscular innervation of either the body or base specimens of the decentralized bladder. While adrenergic muscular innervation of both regions was similar to normal in short-term samples of the same specimens, there was an obvious adrenergic hyperinnervation of both in the long-term samples. By contrast, there was an obvious, early-developing, and persistent hypoinnervation by both cholinergic and adrenergic nerves in both the body and base regions after postganglionic “denervation.” It is concluded that histochemistry (1) has no value in detecting either short- or long-term changes in cholinergic muscular innervation of the decentralized bladder and (2) is useful for revealing both short- and long-term cholinergic and adrenergic hypoinnervation of the “denervated,” as well as long-term adrenergic hyperinnervation of the decentralized vesical muscularis.     

Intrinsic neuromuscular defects in the neurogenic bladder. III. Transjunctional,short- and long-term ultrastructural changes in muscle cells of the decentralized feline bladder base following unilateral sacral ventral rhizotomy

We have shown previously that vesicourethral decentralization by unilateral sacral ventral rhizotomy in the cat results in sequential degenerative and regenerative ultrastructural changes in intrinsic nerves of the bladder base. The present study in the same model provides evidence that these neural changes are associated with both degenerative and regenerative ultrastructural changes in smooth muscle cells of the feline base, representing the first example of transjunctional postdecentralization change in an effector tissue to be documented in a mammalian autonomically innervated smooth muscle system. The nature of the observed changes in the sarcoplasm, sarcoplasmic organelles, sarcolemma, and cell-to-cell relationships will be discusssed, and their physiologic significance as well as implications in the phenomenon of neurogenic muscular supersensitivity will be presented.     

Intrinsic neuromuscular defects in the neurogenic bladder: IX. Effects of combined parasympathetic decentralization and hypogastric neurectomy on neuromuscular ultrastructure of the feline bladder base

Previous studies have shown that unilateral decentralization of the feline bladder by sacral ventral rhizotomy results in ultrastructural neuromuscular changes in the bladder base. In the present study, the neuromuscular ultrastructure of the feline bladder base was studied in twenty sexually mature adult male cats 1–4 and 6–10 weeks following bilateral sacral rhizotomy, alone and combined with hypogastric neurectomy. The former procedure resulted in initial degeneration of cholinergic axons with loss of their neuroeffector junctions, but the adrenergic axons and neuroeffector junctions were preserved; within 6–10 weeks, the bladder base became re-innervated by regenerated cholinergic, increased adrenergic, and abundant probable copeptidergic axons. In contrast, combined sacral rhizotomy and hypogastric neurectomy resulted in persistent (up to 10 weeks) degeneration of both cholinergic and adrenergic axons with loss of their neuroeffector junctions, and no adrenergic hyperinnervation, with only a modest increase in probable copeptidergic axons in long-term samples. Our observations indicate that intact efferent sympathetic pathways are essential for eventual cholinergic re-innervation and adrenergic hyperinnervation of the decentralized bladder and suggest that the latter is an important factor in the development of vesical hypertonicity in lower-motor neuron lesions. On this basis, we introduce the concept that two functional types of the lower motor neurogenic bladder–i.e., the hypertonic with intact and the normotonic with injured efferent sympathetic pathways–may be distinguishable morphologically by ultrastructural study of the vesical muscularis.     

Detection of toxigenic Escherichia coli using biotin-labelled DNA probes following enzymatic amplification of the heat labile toxin gene

Several types of DNA probes labelled with biotin were compared for their sensitivity to detect the heat labile toxin (LT) gene in toxigenic Escherichia coli. In addition, a procedure was developed for enzymatically amplifying LT gene sequences in toxigenic E. coli. Probes were labelled with biotinylated nucleotides by either nick translation; 3' tailing; primer extension of probe DNA cloned into bacteriophage M13; sandwich hybridization; or oligolabelling of isolated DNA fragments. A single stranded probe consisting of a DNA fragment from the LT gene cloned into the bacteriophage M13mp18 and detected by hybridization to oligolabelled biotinylated M13mp18 RF DNA in a sandwich hybridization was able to detect as little as 10 pg of toxin gene DNA. Cloned LT gene DNA was serially diluted and amplified enzymatically using synthetic oligonucleotide primers. Amplified DNA was detected using biotin-labelled M13-based probes. As little as 1 fg of LT DNA could be amplified to detectable levels by this method. Experiments with LT+ bacteria resulted in the detection of as few as 1000 bacteria. The combination of enzymatic amplification coupled with M13-based DNA probes provides a highly sensitive tool for detecting pathogenic microorganisms.     

High branched-chain alpha-keto acid intake, branched-chain alpha-keto acid dehydrogenase activity, and plasma and brain amino acid and plasma keto acid concentrations in rats

Diets containing high quantities of individual branched-chain alpha-keto acids (BCKAs) or a combination of BCKAs as used for treatment of renal disease were fed to rats. When the diet contained a single BCKA, its concentration was high in plasma and the concentration of its corresponding amino acid was high in plasma and brain. Liver BCKA dehydrogenase (BCKD) was 42% active in control rats. Consumption of diets containing 0.38 mol/kg diet of alpha-ketoisocaproate (KIC), alpha-keto-beta-methylvalerate (KMV), or alpha-ketoisovalerate (KIV) resulted in complete activation of liver BCKD. Consumption of the diet containing the combination of BCKAs increased basal BCKD activity of liver twofold. Muscle BCKD was activated after feeding the KIV diet (2-fold), the KIC diet (3-fold), and the KMV diet (15-fold). Total BCKD activity of liver and muscle was unaffected by dietary treatments. Activation of liver and muscle BCKD by dietary BCKA is consistent with their ability to inhibit BCKD kinase in vitro.     

Glycosylated fetal hemoglobin. Correlation with hyperglycemia and birth weight in infants of diabetic mothers

Glycosylated fetal hemoglobin levels were measured in umbilical cord blood of normal infants and infants of diabetic mothers. The glycosylated fraction proved to be a stable compound; its level remained unchanged over a 19-day period. Exposure of fetal and adult hemoglobins to the same concentrations of glucose in vitro resulted in similar levels of glycosylated hemoglobins, suggesting that both types of hemoglobin are about equally reactive with glucose. Levels of glycosylated hemoglobin were significantly increased above normal in umbilical cord blood of infants of both Class A and Class B diabetic mothers. A significant relationship was found between macrosomia, reflected in birth weight ratios, and glycosylated hemoglobin from fetal erythrocytes in infants of diabetic mothers. While these data are consistent with the conclusion that glycosylated fetal hemoglobin levels are a function of fetal blood glucose concentrations in utero during the 2 mo before delivery, it is not known whether the glycosylated hemoglobin contributes to the abnormalities other than macrosomia found in infants of diabetic mothers.     

Transformation reactions of progesterone by different species of Streptomyces

A systematic study of transformation reactions of the genus Streptomyces with respect to the progesterone molecule was undertaken. The types of transformation reactions by different Streptomyces species were evaluated from the point of view of taxonomy. The isolates tested were divided according to the transformation types into six groups: (1) a group of species transform progesterone to 16α-hydroxyprogesterone; (2) a group of species transform progesterone to 6β-hydroxyprogesterone; (3) a group of species transform progesterone to 6β, 11α-dihydroxyprogesterone; (4) a group of species dehydrogenate progesterone in C_{1 - 2} position; (5) a group of species transform progesterone to 3 derivatives namely 6β-hydroxyprogesterone, 6β, 11α-dihydroxyprogesterone and dehydrogenation in position C_{1 - 2}; (6) a group of species with no capacity to transform progesterone into another steroid derivative. From the point of view of Streptomyces classification, the transformation reaction of progesterone fulfil all the requirements of taxonomic feature of Streptomyces. These appear to be specific properties and common to all the strains of individual Streptomyces species tested.