Inge K. Genefke

Early epidermal changes in heat- and electrically injured pig skin. II. An electron microscopic study☆

In order to find methods applicable for disclosing electrical torture, pig skin was exposed to heat and electricity under controlled circumstances. Biopsies for electron microscopy were obtained immediately after exposure. In heat lesions the nuclei were slightly distorted, sometimes with broken nuclear membranes. The tonofilaments were clumped, intracellular oedema was present and cell membranes were ruptured between desmosomes. In electrical lesions the nuclei were usually enlarged with strongly condensed chromatin. Some nuclei were composed of fine, evenly dispersed granular material. The cytoplasm appeared homogeneous, in large magnification finely granular. Cell borders could sometimes be identified located in situ. In the stratum corneum, which appeared normal in heat lesions, single or several cells or large areas had an electron-dense appearance. The difference in ultrastructure of heat and electrical lesions makes it probable that electricity has a specific action on epidermal cells.

Tracing the use of electrical torture.

With the aim of being able to trace skin sequelae to electrical torture, an interdisciplinary group of scientists (the “electrical group” of Anti-Torture Research, ATR) has performed controlled morphological studies on skin biopsies from experiments with fully anesthetized pigs. “Vesicular nuclei” in epidermis and a characteristic pattern of collagen calcification in dermis were found to be typical of electrical damage. These alterations were produced by alternating current as well as by direct current

Electrically-induced collagen calcification in pig skin. A histopathologic and histochemical study

Deposition of calcium salts on collagen fibres in skin of fully anaesthetized pigs was induced by exposure to direct current (d.c.). In biopsies obtained from cathode areas successively from day 1 to day 7 after exposure the histopathologic and histochemical changes before and after the initial deposition of calcium salts have been examined. For comparison skin sites with intradermal injected calcium hydroxyapatite crystals were studied in addition. Small areas of calcified collagen and elastic fibres were noted in viable tissue 2 days after d.c. exposure. In succeeding days the calcified areas enlarged with new deposits always more superficial and closer to the epidermis than the original calcium deposits. Preconditions for calcification appear to be (1) a pH change in basic direction and/or the electrochemical processes specific to the cathode area and (2) a viable tissue. Elastic fibres appear to have a lower calcification threshold than collagen fibres. A positive staining for glycoproteins (PAS) and glycosaminoglycans (alcian blue pH 2.5) was noted in the calcified collagen fibres simultaneously with the calcification. In succeeding days the intensity of the staining reactions increased. Whether changes in the glycoproteins, collagen and its intimately bound glycosaminoglycans precede the calcification or the staining reactions develop secondarily to this deposition is not known. However, seven days after intradermal injections of Ca-apatite crystals in pig skin small and large crystals were observed ultrastructurally without any relation to collagen fibrils, but the calcified tissue presented a positive PAS and alcian blue reaction from day 2. Thus the PAS and alcian blue stainings in this model develop secondary to the deposition of calcium salts.

Ultrastructural changes in dermal pig skin after exposure to heat and electric energy and acid and basic solutions

In order to describe the ultrastructure of the histopathological changes in dermis after exposure to electrical energy, heat energy and acid and basic solutions the skin of fully anaesthetized Danish Landrace pigs were exposed to direct current, heat (80 degrees C and 450 degrees C) and acid and basic solutions. Biopsies were obtained immediately after the exposure from all types of injury. Biopsies from the cathode areas biopsies were also taken on day 1 and day 2.5 in order to describe the initial calcium deposits. Homogeneous collagen fibres without any birefringence from heat exposed areas were ultrastructurally composed of filamentous materials. Collagen fibres with fine densely spaced cross-striation from cathode areas and areas exposed to basic solutions were shown ultrastructurally to consist of parallelly arranged collagen fibrils with regular waves. It is concluded that the cross-striation of the collagen fibres observed in polarized light are due to a periodic change in the orientation of the fibres seen as waves of the fibres. The ultrastructure of dermal cells were similar to that of epidermal cells following the different types of influence. Characteristically the nuclei were condensed following heat and more electron-lucent following direct current (d.c.) and acid and basic solutions. In cathode areas and areas influenced by basic solutions the electron-lucent nuclei contained fine fibrils. The ultrastructural study supports the suggestion from light microscopic studies that the morphology of anode and cathode lesions shows resemblance to acid induced and basic induced lesions, respectively. Apatite crystals were observed on day 2.5 at the periphery of the collagen fibrils and in the matrix of elastic fibres.

The active uptake of 5-hydroxytryptamine in rat and human blood platelets under the influence of lithium in vivo and in vitro.

Several investigators relate the concentration and turnover of 5-hydroxytryptamine (5-HT) in brain to affective disorders (Murphy et al. (1969), Dencker et al. (1966), Roos & Sj6strom (1969), Mendels et al. (1971), Lapin & Oxenkrug (1969) ). As lithium is today extensively used in the treatment of manicdepressive illness (Bamtrup et al. (1970)), we found it of interest to investigate the effect of lithium on a part of 5-HT metabolism. In brain, 5-HT is found in intracellular storage organelles in nerve-endings. In blood platelets the 5-HT is also localized in intracellular organelles ( D a Prada & Pletscher (1969), Tranzer et al. (1966)). An active uptake mechanism capable of transporting 5-HT into the cells has been demonstrated both in blood platelets and in brain (Pletscher (1968), Ross & Renyi (1967), Blackburn et al. ( 1967) ). The similarities between 5-HT in brain and platelets have led us to investigate, as proposed by Page (1968) and Ahtee (1968), the effect of the drug on the active transport of 5-HT into blood platelets, as a useful model system. We have examined platelets from rats treated with lithium as well as the transport of 5-HT in rat and human thrombocytes, when lithium was added to the incubation medium.

THE CONCENTRATION OF 5‐HYDROXYTRYPTAMINE (5‐HT) IN HYPOTHALAMUS, GREY AND WHITE BRAIN SUBSTANCE IN THE RAT AFTER PROLONGED ORAL LITHIUM ADMINISTRATION

The thrombocyte is considered a useful model for the monoamine containing nerve endings in brain. In a previous paper, lithium was found to inhibit the in uitro uptake of 5-hydroxytryptamine (5-HT) in rat and human blood platelets, while lithium administration for 1-8 weeks did not change the 5-HT content of rat platelets or the active 5-HT uptake in the platelets (Genefke & Langgird (1971)). After administration of lithium salts, the lithium ion is not equally distributed in the rat brain. Following prolonged administration, Edelfors & Gclthgen (1971) demonstrated that the concentration of lithium is two to four times greater in the hypothalamic region than in other parts of the brain. We therefore found it of interest to measure the content of 5-HT in different regions of the rat brain after prolonged peroral administration of lithium. The 5-HT content was determined in three parts of the brain: hypothalamus, grey substance from the frontal cerebral cortex, and white substance from the area near capsula interna.

METHODOLOGICAL INVESTIGATIONS ON THE IN VITRO UPTAKE OF 5‐HYDROXYTRYPTAMIN BY HUMAN BLOOD PLATELETS

Blood platelets and monoamine-containing nerve endings of the brain share anatomical and functional properties, particularly with regard to active uptake, storage, and release of 5-hydroxytryptamin (serotonin). Thus experimentally induced changes in the serotonin metabolism of blood platelets may reflect changes occurring intraneuronally (Page ( 1968), Paasonen ( 1968), Murphy ( 1969) ). In a series of investigations on the effect of drugs used in manic-depressive illness, e. g. lithium, on brain serotonin metabolism, we were therefore interested in examining also the effect of these drugs upon blood platelets in vit 7 0 . However, with this particular aim of the study, it was felt that some of the usually applied analytical procedures for examination of blood platelets might lead to erroneous conclusions. We were, in this respect, particularly concerned about the use of Ca++and Mg++ -removing anticoagulants. The present study was undertaken to establish the significance, for this particular type of investigation, of some analytical details, i. e. type of incubation medium, incubation period, concentration of serotonin and pH of the incubation medium, and the use of anticoagulants.