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Morphological and functional identifications of catfish retinal neurons. II. Morphological identification.

In this study the morphological origins of the responses from the catfish retinal neurons evoked by step inputs were determined by injecting intracellularly a dye, Procion yellow. A method was devised to view the dye-injected neurons in flat mount to study their dendritic expansion; later the same neurons could be sectioned radially to locate the levels of their somata or dendritic expansion. The results of this study show the inherent danger of identifying dye-injected neurons only in a radial or tangential view. Bipolar cells could be identified functionally without any ambiguity by changing widely the stimulus parameters, because the stimulation of their receptive-field center and surround gave rise to responses of opposing polarity. We found no exception to this rule. The neurons in the proximal layers produced a large variety of responses which could not be segregated into two such classes as the amacrine and ganglion cells. In this part II they were classified into three broad categories: neurons giving rise to sustained, transient, and spiking responses. The demarcation among the three types, morphologywise and functionwise, was vague and not well established. The sustained responses were recoreded from the starburst and spaghetti neurons (part I (9)) which correspond to Ramón y Cajal's (2) amacrine cells. The transient responses, whose patterns were largely invariant of the changes in the stimulus parameters, were recorded from a class of neurons with spindle-shaped somata in the INL. We do not know whether they had axons or not, but we will not be surprised if a future study defines them as a class of ganglion cells. Responses with or without spike discharges were recorded from a class of neurons which were identified as ganglion cells. Observations made on a large number of Procion-injected neurons in both flat-mount preparations and radial sections show that finer dendritic arborizations were not seen in the dye-injected neurons although the presence of such branches was proved in the Golgi preparations. Probably this was due to the weak contrast of the Procion-injected cell against the tissue background, rather than the failure of the dye to diffuse into finer branches. We recognize the severe difficulty involved in the traditional approach of identifying a class of neurons based on typical but subjectively selected functional and structural samples. Neurons have to be classified statistically according to their (quantitative) parameters. (cont'd)

Animals↗

Identification of a macromolecular factor in the ileum which binds intrinsic factor and immunologic identification of intrinsic factor in ileal extracts.

The precipitate which resulted when (57)CoB(12) bound to normal human gastric juice was subjected to a 15% concentration of Na(2)SO(4) contained virtually no radioactivity. However, after in vivo incubation of the gastric juice-(57)CoB(12) mixture in the distal ileum of the guinea pig, the dialyzed extract of the washed mucosa contained a fraction of (57)CoB(12) which was precipitated at 15% Na(2)SO(4). In addition, in vitro incubation of gastric juice-(57)CoB(12) with an extract of the ileal mucosa or brush border membranes also resulted in the formation of a 15% Na(2)SO(4)-insoluble fraction which contained (57)CoB(12). The formation of this (57)CoB(12)-containing insoluble fraction did not occur or was diminished by (a) addition of an excess of B(12)-free normal human gastric juice. (b) reducing the incubation pH to 2, (c) incubating the mixture at 4 degrees C, (d) pretreating the ileal extract at 56 degrees C for 30 min, (e) incubating the reaction in sodium EDTA but not calcium EDTA, (f) incubating gastric juice-(57)CoB(12) with an extract of jejunal mucosa. Sephadex gel filtration was used to demonstrate that the factor in the ileal extract which reacted with the gastric juice-(57)CoB(12) filtered through G-100 and G-200 columns in the excluded volume. When the ileal extract obtained after in vivo incubation with gastric juice-(57)CoB(12) was subjected to starch gel electrophoresis one peak of radioactivity remained at the origin and another moved anodally. Eluates of each peak reacted with anti-intrinsic factor antibody indicating that at least the immunologically reacting portion of the intrinsic factor molecule was present in two fractions with different electrophoretic mobility.These studies indicate that immunologically intact intrinsic factor can be extracted from the ileum after in vivo incubation with gastric juice-(57)CoB(12), and that a macromolecular factor is present in the distal ileal mucosa which binds intrinsic factor both in vitro and in vivo, changing its solubility and electrophoretic properties. It is suggested that this ileal binding factor is the previously postulated intestinal receptor for intrinsic factor.

Animals↗

[Isolation and identification of Cryptosporidium from various animals in Korea. II. Identification of Cryptosporidium muris from mice].

Each of SPF mice(Scl: ICR strain, 3-week-old males) was inoculated with 5 x 10(4) oocysts of Cryptosporidium by stomach tube. The oocysts were large type one which was previously isolated from Korean mice, and passaged in 3-week-old SPF mice. The patterns of oocyst discharge were monitored daily, and in order to observe the ultrastructure of developmental stages the stomach of the mice was examined by transmission electron microscopy (TEM) at 4 weeks post-inoculation. The prepatent period for 6 mice was 5.6 days post-inoculation on the average, and the patent period was 63.2 days. The number of oocysts discharged per day from the mice reached peak on day 36.6 post-inoculation on the average. A large number of oocysts were found in fecal samples obtained from inoculated mice on days 30-50 post-inoculation. C. muris was larger than C. parvum at almost every developmental stages, the size difference being 1.4 times in oocysts, 2.4 times in sporozoites, 1.6 times in merozoites, and 1.5 times in microgametes. The ultrastructural features of the attachment site of C. muris to the mucus cells were remarkably different from those of C. parvum and its closely related species. The anterior projection of the protozoa (C. muris), the outer aspect of which was surrounded by a thick filamentous process of the host cell, has not been reported at any developmental stages of C. parvum or its closely related species. The size of the oocysts of strain RN 66 was larger than that of Korean mice origin. The above results reveal that the large type Cryptosporidium of Korean mice origin is identified as Cryptosporidium muris and this type was named as C. muris (strain MCR).

Animals↗

[Isolation and identification of Cryptosporidium from various animals in Korea. III. Identification of Cryptosporidium baileyi from Korean chicken].

Each of SPF chicken (Hi-Line strain, 2-day-old males) was inoculated with 2.5 or 5 x 10(4) oocysts by stomach tube. The oocyst was the medium type of Cryptosporidium previously isolated from Korean chicken origin, and passed in 2-day-old SPF chicken. The patterns of oocyst discharge were monitored daily, and in order to observe the ultrastructure of the developmental stages, the bursa of Fabricius of the chicken was examined by transmission electron microscopy (TEM) on the 12th day postinoculation. The prepatent period for 8 chicken was 5.9 days postinoculation on the average, and the patent period was 12.9 days. The number of oocysts discharged per day for the chicken was reached peak on day 12 postinoculation on the average. A large number of oocysts was found in fecal samples obtained from inoculated chicken on days 8-14 postinoculation. The ultrastructural feature of almost every developmental stage of the medium type from chicken was very similar to that of Cryptosporidium previously isolated from mammalia including human and birds except for the attachment site of C. muris to the mucus cell from mammalia, but dimension of the oocysts from fecal samples of the medium type was different from those of C. meleagridis and mammalia origin. The above results reveal that the medium type of Cryptosporidium of Korean chicken origin is identified as Cryptosporidium baileyi.

Animals↗

Double-gradient denaturing gradient gel electrophoresis assay for identification of L-ferritin iron-responsive element mutations responsible for hereditary hyperferritinemia-cataract syndrome: identification of the new mutation C14G.

BACKGROUND: Hereditary hyperferritinemia-cataract syndrome is an autosomic dominant disorder caused by heterogeneous mutations on the iron-responsive element (IRE) of ferritin L-chain mRNA. The mutations described to date were identified by direct sequencing of DNA from probands with hyperferritinemia often associated to bilateral cataracts. A direct genetic approach on a large population is useful to recognize polymorphisms in the DNA region and the prevalence of mutations associated with minor increases in serum ferritin and subclinical cataracts. We developed a rapid DNA scanning technique to detect mutations in a single electrophoretic analysis. METHODS: The double-gradient denaturing gradient gel electrophoresis (DG-DGGE) method consisted of PCR amplification of the target genomic DNA with GC-clamped oligonucleotides. The sequence encoded the 5' untranslated flanking region of ferritin L-chain mRNA, which includes an IRE stem-loop structure. The product was subjected to DG-DGGE (8.5-15% polyacrylamide and 50-95% denaturant) to separate the homo- and heteroduplexes. RESULTS: The method clearly identified all eight accessible mutations, including C-G transversions, which are the most difficult to detect. The method was applied to scan DNA samples from 50 healthy subjects and from 230 subjects with serum ferritin >400 microg/L. The new mutation G14C was identified. CONCLUSIONS: The DG-DGGE method detects all the mutations in the L-ferritin IRE sequence, is rapid and economical, and can be applied to scan large populations. The first population study indicated that the mutations are rare and may involve regions of the IRE structure not yet characterized.

Base Sequence↗

Identification of a novel cell type in peripheral lymphoid organs of mice. IV. Identification and distribution in mouse spleen.

White pulp nodules of mouse spleen contain a minor population of cells with morphologic features that are identical to those of dendritic cells, a cell type recently described in vitro. They have characteristic large, irregularly shaped nuclei with distinctive chromatin patterns and small nucleoli. The cytoplasm is extended in processes that contain relatively few organelles. These presumptive dendritic cells can be distinguished from other cell types that are known to exist in spleen including those that have irregular or branching cell shapes. In particular, dendritic cells do not contain the large number of lysosomes seen in phagocytes, and do not actively interiorize intravenously administered colloidal thorium dioxide particles. They also lack the well developed secretory apparatus (rough endoplasmic reticulum and Golgi zone) and microfilament bundles that are noted in connective tissue cells. These morphologic observations, combined with previous in vitro work, substantiate the existence of a novel class of cells in mouse lymphoid organs.

Animals↗

[Identification of the sinus potential in man by endocavitary electrocardiography. Identification criteria, preliminary results].

The potential of the sinus node was recorded in 24 patients by endocavitary electrocardiography using the same equipment as for usual electrophysiological investigation. The sinus node potential is a slow wave which starts progressively, has a rounded peak and precedes the endocavitary atrial activation and the surface P wave, and is relayed at an acute angle to the high amplitude deflection of atrial depolarisation. In some specially selected recordings it is easy to confirm the sinus origin of this slow wave where its preatrial position may be defined by the fixed relation of the S-P interval and by the variability of the T-S interval. Usually it is necessary to use technical manoeuvres (vagal stimulation, premature atrial stimulation) to demonstrate the preatrial behaviour of a potential thought to be of sinus origin. The variability of the sinus potential and the apparent sinoatrial conduction time, is suggestive of a physiological instability of the intrasinus pacemaker. The sinoatrial conduction time measured directly does not seem to exceed 140 ms in normal subjects.

Arrhythmias, Cardiac↗