Baker's Acid Research Articles

The Effects of Single Doses of Pilocarpine and of Histamine upon the Granules of Oxyntic Cells of Rats, with Special Reference to their Phospholipine Content

ABSTRACT New data are presented on the morphology and cytochemistry of the granules of the oxyntic cells of the rat’s stomach in continuation of work previously reported (Menzies, 1949). The effects of single injections of pilocarpine and of histamine on the granules were investigated, with particular reference to their phospholipine content as shown by Baker’s acid haematein method. Pilocarpine causes a marked enlargement of the granules with loss of phospholipine from the central part of many of the larger granules, and complete loss of phospholipine from others. The granules that have lost their phospholipine probably still contain lipoid as shown by sudan black. Pilocarpine also causes marked aggregation of the granules. Histamine causes a less marked increase in granule size and no observable loss of phospholipine, but some of the granules became elongated, an effect that can be seen with the phase-contrast microscope, and there is a tendency for them to recede to the peripheral parts of the cells associated with an enlargement of the intracellular canals. Both pilocarpine and histamine result in some enlargement of the oxyntic cells, the effect being more marked with pilocarpine. Neither pilocarpine nor histamine alters the size or shape of the non-lipoid moiety of the granules. Supravital colouring with neutral red shows a few droplets in oxyntic cells in the bases of the gastric glands. These enlarge slightly after histamine and greatly after pilocarpine, and they are always spherical. It has not yet been possible to decide as to their exact nature.

Cytochemical Reactions of Bovine Spermatozoa and Seminal Plasma

Summary By using accepted cytochemical techniques the following reactions were observed in bovine spermatozoa: Acid phosphatase was present throughout with a slight reaction in the equatorial segment when sodium glycerophosphate was used as a substrate at pH 5.0. Alkaline phosphatase was present, with the exception of the neck, when the same substrate was employed at pH 9.0. Enzymatic activity was found in the neck, middle-piece and tail when adenosine triphosphate was used at pH 5.0. The occurrence of an enzyme acting upon hexose diphosphate (fructose-l,6-diphosphate) at pH 9.0 was noted in the head, neck, middle-piece and tail. Gram's stain and the Feulgen reaction were positive throughout the head, indicating the presence of both ribonucleic and desoxy-ribonucleic acids. Metachromatic staining was not obtained with toluidin blue, indicating the absence of mucopolysaccarides. Fat, including phospholipid, was localized in the middle-piece. The acrosome was characterized by a slight positive reaction with the periodic acid-Schiff reaction which was not due to glycogen, glycolipid or mucopolysaccharide. Furthermore, the protoplasmic droplet which occasionally was present on the tail gave an intense reaction with the periodic acid-Schiff procedure. Seminal plasma gave the following reactions: Acid phosphatase was positive with sodium glycerophosphate as a substrate at pH 5.0 and alkaline phosphatase was characterized by a weak response with hexose diphosphate (fructose-1,6-diphosphate) at pH 9.0. Gram's stain and toluidin blue gave no reactions. Baker's acid hematein test for phospholipid was negative, but the control preparations were characterized by a bluish-grey stippling and a positive test was obtained with Sudan black B. No reaction was observed with Bodian's protargol method. A slight positive reaction was obtained with the periodic acid-Schiff reaction.

Observations on the Chemical Composition of Myelin and the Smallest Size of Myelinated Nerve-Fibres in the Central Nervous System

Baker's (1946) acid haematein and pyridine-extraction control tests, claimed to be specific for phospholipines (Baker, 1947), have been applied to various parts of the central nervous system of rats and man. The sudan black method for the detection of lipoids and the nile blue method for the staining of acidic lipoids have also been used. The findings are in agreement with older statements in the literature that myelin contains a considerable amount of phospholipines. It was impossible to determine whether galactolipines or neutral lipoids are also present. In the acid haematein-stained sections finer fibres were seen than when other stains for myelin sheaths are employed. Fibres with a diameter of 0.5 µ or even somewhat less were stained in various parts of the central nervous system of rats. It is regarded as probable from these findings that fibres down to 0.5 µ or even smaller possess a lipoid investment. These observations lend support to the now commonly accepted view that the distinction between myelinated and so-called unmyelinated fibres is arbitrary. Some observations are made, however, which indicate that the presence of truly unmyelinated fibres cannot be excluded.

An Examination of Baker's Acid Haematein Test for Phospholipines

1. Baker's acid haematein test for phospholipines is specific provided that only a definite positively result is considered. Very pale blues and greys may be caused by other lipoids, which if present in very large masses may possibly show medium to dark blue granules but will not be coloured all through. 2. The mechanism of the test appears to be as follows: (a) Phospholipine is not fixed by formal-calcium but is restrained from passing into solution by the calcium ions, which play no other part. (b) Phospholipine combines readily with chromium from the mordanting fluid, and is then rendered insoluble and mordanted. Other substances, acidic and usually containing phosphorus, are mordanted as well. (c) On staining, blue and brown colorations are formed; in both cases the dye attaches itself to the chromium in the various substrates. (d) On differentiation, some browns and most blues, particularly those with phosphoric substrates, remain nearly fast, but most browns and the weak blues of certain lipoids (not phospholipines) are greatly reduced or removed entirely. The period of differentiation must not be shortened. (e) Blue-staining lipoids (phospholipines) are distinguished from other blue-staining substances by an extraction with the lipoid solvent pyridine, after special fixation. The other substances, and any bound lipoid not removable with pyridine, remain. 3. Since the specificity of the test depends on the relatively greater affinity of phospholipines among lipoids for the mordant, the period of chroming must not be lengthened. 4. One reason why some substances are coloured after pyridine extraction but not after acid haematein is that in the former case they are precipitated and so concentrated; in the latter they are not. This is not a general explanation for the whole class of such substances.