Can membrane excitation be described without a membrane capacity?

A simplified model of excitation is introduced in which the membrane capacity is ignored. It is shown that: (1) Threshold, action potentials, and strength-duration relation can be reproduced by a membrane without a capacity, even for a very simplified model. (2) The delayed build up of the sodium conductance can mimic a membrane capacity. (3) A constant potential stimulus can be used to reveal the influence of the membrane capacity, eventually combined with a feed back mechanism which reduces the effect of the capacity. (4) The effect of the membrane capacity depends on the ratio between the membrane time constant and the time constant for the fast conductance changes.     

Mucosal epithelial cells and Langerhans cells are targets for infection by the immunosuppressive type D retrovirus simian AIDS retrovirus serotype 1

Simian acquired immune deficiency syndrome (SAIDS) caused by the type D retrovirus SRV-1 results in opportunistic infections and a spectrum of oral lesions similar to those seen in humans with AIDS. To better understand the pathogenesis of these oral lesions we have retrospectively examined the oral mucosa from ten rhesus monkeys that died with SAIDS and prospectively examined the oral mucosa of ten additional animals inoculated with SRV-1 to determine at what time, and in what cells SRV-1 infection of the oral mucosa occurs. Using single and double label immunohistologic techniques, and electron microscopy we detected SRV-1 in clusters of oral epithelial cells and rare Langerhans cells as early as 1 month postinoculation.     

The influence of fixation on the relative amount of cytoplasmic ribosomes in mouse epidermal basal keratinocytes

The nature and significance of so-called dark keratinocytes in the epidermis during chemical carcinogenesis is still a matter of concern and debate. Based on ultrastructural observations it has been suggested that dark cells most often are shrunken cells. Reports on skin carcinogenesis, however, claim that dark cells are a sign of ongoing tumor promotion and represent those stem cells in the epidermis from which the tumors originate. It is therefore important to find out whether these cells are simply injured and shrunken cells, or vital cells of great importance for carcinogenesis. Dark cells are assumed to be rich in ribosomes. There is evidence, however, that the observed number of dark cells is highly dependent on tissue fixation. In the present ultrastructural study, morphometric methods were used to compare the effects of two different fixation procedures on the amount of cytoplasmic ribosomes in dark cells from both untreated and carcinogen-treated hairless mouse epidermis. The results show that the ultrastructural features of both dark and clear cells vary considerably with different fixation procedures. In acetone-treated controls typical dark cells are only observed when the fixative has a lower osmotic activity than the plasma. With iso-osmolal fixation typical dark cells are not observed. After an abortive two-stage carcinogenesis treatment, in which a single application of 9,10-dimethyl-l,2-benzanthracene (DMBA) in acetone was followed by a single application of 12-O-tetradecanoyl-13-acetate (TPA) in acetone, signs of cell injury could be found after both fixation procedures. With DMBA/TPA and hypo-osmolal fixation the number of dark cells seemed to increase, whereas only signs of cell injury with occurrence of some heavily altered “clear cells” dominated the picture with iso-osmolal fixation. Morphometry showed that both the numerical and the volumetric densities of cytoplasmic ribosomes in basal keratinocytes varied most significantly with the fixation procedure used. The cytoplasmic volumes did not vary in a way that could explain these differences. One might therefore assume that the number of ribosomes depends on the fixative. Large swelling artifacts occurred when a fixative with low osmotic activity was used, leading to compression of neighboring cells. Hence, an increased ribosomal density reported previously in dark cells is probably related to such cell volume artifacts and does not reflect an actually increased quantity of ribosomes. With both fixation procedures, a single application of DMBA followed by one of TPA appeared to produce an increased number of ribosomes in basal keratinocytes. When hypo-osmolal fixation was used, however, treatment with DMBA/TPA did not influence the cytoplasmic volume or the numerical density of ribosomes, in dark cells. This might indicate that so-called dark keratinocytes following DMBA/TPA treatment are functionally inactive cells that appear more vulnerable than active cells to compression during hypo-osmolal fixation.     

Myocardial S-adenosylhomocysteine hydrolase is important for adenosine production during normoxia

The coronary vasodilator adenosine can be formed in the heart by breakdown of AMP or S-adenosylhomocysteine (SAdoHcy). The purpose of this study was to get insight into the relative importance of these routes of adenosine formation in both the normoxic and the ischemic heart. A novel HPLC method was used to determine myocardial adenosine and SAdoHcy. Accumulation of SAdoHcy was induced in isolated rat hearts by perfusion with L-homocysteine thiolactone or L-homocysteine. The release of adenosine, inosine, hypoxanthine, xanthine and uric acid was determined. Additional in vitro experiments were performed to determine the kinetic parameters of S-adenosylhomocysteine hydrolase. During normoxia the thiolactone caused a concentration-dependent increase in SAdoHcy. At 2000 microM of the thiolactone an SAdoHcy accumulation of 0.49 nmol/min per g wet weight was found during normoxia. L-Homocysteine (200 microM) caused an increase of 0.37 and 4.17 nmol SAdoHcy/min per g wet weight during normoxia and ischemia, respectively. The adenosine concentration in ischemic hearts was significantly lower when homocysteine was infused (6.2 vs. 11.5 nmol/g; P less than 0.05). Purine release was increased 4-fold during ischemia. The Km for hydrolysis of SAdoHcy was about 12 microM. At in vitro conditions favoring near-maximal SAdoHcy synthesis (72 microM adenosine, 1.8 mM homocysteine), the synthesis rate in homogenates was 10 nmol/min per g wet weight. From the combined in vitro and perfusion studies, we conclude that S-adenosylhomocysteine hydrolase can contribute significantly to adenosine production in normoxic rat heart, but not during ischemia.     

Taxonomic studies of the genus Hypoxis in East Africa

East African material of the genus Hypoxis L. has preliminarily been divided into the heterogenous, probably apomictic H. obtusa Burch- complex (2n = 40–50, ca. 75, 76, ca. 85, >86, ca. 92, ca. 98, ca. 108, 130–135, 160–200) and 5 rather homogenous species: H. angustifolia Lam. (2n = 14, 28), H. goetzei Harms (2n = ca. 62), H. kilimanjarica Bak., H. malosana Bak. (2n = 14) and H. macrocarpa Holt & Staubo sp. nov. H. kilimanjarica is divided into ssp. kilimanjarica and ssp. prostrata Holt & Staubo ssp. nov.     

Myocardial S-adenosylhomocysteine hydrolase is important for adenosine production during normoxia

(1) The coronary vasodilator adenosine can be formed in the heart by breakdown of AMP or S-adenosylhomocysteine (SAdoHcy). The purpose of this study was to get insight into the relative importance of these routes of adenosine formation in both the normoxic and the ischemic heart. (2) A novel HPLC method was used to determine myocardial adenosine and SAdoHcy. Accumulation of SAdoHcy was induced in isolated rat hearts by perfusion with L-homocysteine thiolactone or L-homocysteine. The release of adenosine, inosine, hypoxanthine, xanthine and uric acid was determined. Additional in vitro experiments were performed to determine the kinteic parameters of S-adenosylhomocysteine hydrolase. (3) During normoxia the thiolactone caused a concentration-dependent increase in SAdoHcy. At 2000 μM of the thiolactone an SAdoHcy accumulation of 0.49 nmol/min per g wet weight was found during normoxia. L-Homocysteine (200 μM) caused an increased of 0.37 and 4.17 nmol SAdony/soc per g wet weight during normaxia and ischemia, respectively. (4) The adenosine concentration in ischemic hearts was significantly lower when homocysteine was infused (6.2 vs. 115 nmol/g; P < 0.05). Purine release was increased 4-fold during ischemia. (5) The K_m for hydrolysis of SAdoHcy was about 12 μM. At in vitro conditions favoring near-maximal SAdoHcy synthesis (72 μM adenosine, 1.8 mM homocysteine), the synthesis rate in homogenates was 10 nmol/min per g wet weight. (6) From the combined in vitro and perfusion studies, we comclude that S-adenosylhomocysteine hydrolase can contribute significantly to adenosine production in normoxic rat heart, but not during ischemia.     

Myocardial xanthine oxidase/dehydrogenase

High-energy phosphates in heart muscle deprived of oxygen are rapidly broken down to purine nucleosides and oxypurines. We studied the role of xanthine oxidase/dehydrogenase (EC 1.2.3.2/EC 1.2.1.37) in this process with novel high-pressure liquid chromatographic techniques. Under various conditions, including ischemia and anoxia, the isolated perfused rat heart released adenosine, inosine and hypoxanthine, and also substantial amounts of xanthine and urate. Allopurinol, an inhibitor of xanthine oxidase, greatly enhanced the release of hypoxanthine. From the purine release we calculated that the rat heart contained about 18 mU xanthine oxidase per g wet weight. Subsequently, we measured a xanthine oxidase activity of 9 mU/g wet wt. in rat-heart homogenate. When endogenous low molecular weight inhibitors were removed by gel-filtration, the activity increased to 31 mU/g wet wt. Rat myocardial xanthine oxidase seems to be present mainly in the dehydrogenase form, which upon storage at -20 degrees C is converted to the oxidase form.     

Lipase-Catalyzed Regioselective Acylation and Deacylation of Glucose Derivatives

In the development of an efficient synthesis of 1-O-decanoyl-2,3,4,6-tetra-O-acetyl-β-D-glucose (β-2) several lipase-based approaches have been explored. Among five immobilized Upases tested, the lipase from Candida antarctica proved particularly efficient for catalyzing selective hydrolysis in the 1-position of 1,2,3,4,6-penta-O-acetyl-β-D-glucose (β-1). Using triethylamine as catalyst, the hydrolysis product 2,3,4,6-tetra-O-acetyl-D-glucose (3) can be esterified with decanoyl chloride to form β-2 selectively, thereby providing an efficient chemo-enzymatic synthesis starting from readily available raw materials. Attempts to produce β-2 from β-1 by lipase-catalyzed interesterification or to esterify 3 with decanoic acid using a lipase as catalyst were unsuccessful. The latter finding was explained by the hemiacetal OH group of glucose being unable to act as nucleophile in the lysis of the lipase acyl-enzyme intermediate. Furthermore, β-2 was found to bee a too bulky substrate to fit into the active site of any of the lipases tested.