Influence of light and darkness on the hypothalamo-hypophysial system of Rana temporaria L.

Summary The influence of a long daily photoperiod and continuous darkness was studied on the hypothalamo-hypophysial system of Rana temporaria. After six weeks of treatment (December-January 1968/1969) the secretory activity of the hypothalamo-hypophysial system of the light-treated animals was enhanced.The amount of aldehyde-fuchsin positive material in the preoptic nucleus increased as a result of the light treatment, and decreased in the median eminence, whereas in the posterior lobe no difference could be observed between the light- and dark-treated animals.The karyometric results showed an increase of the nuclear volume of the cells of the preoptic nucleus after light treatment; small differences were noticed between distinct areas of the preoptic nucleus.Autoradiographic data especially showed that light influenced the hypothalamo-hypophysial system to a high degree. Half an hour after the injection of ³⁵S-cysteine, the incorporation of the isotope in the preoptic nucleus of the light-treated animals was five times as much as in the dark-treated animals. In the median eminence and in the posterior lobe, the autoradiographic results confirmed the results found with regard to the amount of aldehyde-fuchsin positive material. Labelled material is released only from the median eminence and not from the posterior lobe.The author is indebted to Prof. Dr. J. C. van de Kamer and Dr. F. C. G. van de Veerdonk for their interest and helpful encouragement. Thanks are also due to Ir. J. J. Bezem for his assistance in solving the statistical problems and to Mr. H. van Kooten and his staff for making the illustrations. The co-operation of the student Mr. E. Schenk is gratefully acknowledged. Dr. L. Boomgaart was so kind to check and amend the English writing.     

Thymic vascular system of the European common frog,Rana temporaria: a scanning electron-microscopic study of vascular casts

Vascular corrosion casts of the thymus of adult individuals of the European common frog, Rana temporaria, were analysed by scanning electron microscopy. The main arterial vessel, which is derived either from the temporal artery or from the auricular ramus, approaches the central territory of the gland and branches into twigs that, on penetrating the parenchyma, give rise to capillaries. Most of these capillaries run vertically towards the surface of the gland; they either join the superficial capillary plexus or follow this plexus for a variable distance and then run back towards the medulla, forming capillary loops. The former capillaries link with the extensive venous plexus composed of irregular meshes, whereas the latter capillaries join the venules at the cortico-medullary boundary and finally escape into collecting veins on the gland surface. The venous twigs, which join together near the gland, form the main thymic vein, which empties into the external jugular vein. The details of the thymic vasculature of the anuran amphibian, R. temporaria, are compared with those described in mammalian species, viz. the mouse, rat and guinea pig.     

The optic tracts of Rana temporaria and a possible retino-preoptic pathway

Summary The optic tracts and centres of optic terminals of Rana temporaria have been investigated with silver impregnation techniques after unilaterally cutting the optic nerve and autoradiographically after injection of a mixture of tritiated amino acids into the vitreous body of the left eye.The observations on the course of the optic tracts and on the optic terminals in the thalamus and optic tectum are to a great extent in agreement with those of other authors. The probability of a retino-preoptic pathway is supported by the detection of fibres running in dorsal direction in front of the place where the optic nerve penetrates the brain. In horizontal sections these fibres can be seen deviating from the optic tract, covering a short distance in frontal direction and then turning upward under an angle of about 90 degrees. They disappear between the ventral aldehyde-fuchsin positive cells of the preoptic nucleus. The ventral and median parts of the ipsilateral preoptic nucleus contain fragments of degenerated fibres.Autoradiographic data are also in favour of the presence of an ipsilateral retino-preoptic tract. After carefully counting the number of grains over left and right preoptic nucleus, it appeared that over the ventral and median parts of the left preoptic nucleus more grains occur than over the identical areas of the right one.It is a pleasure to acknowledge Prof. Dr. J. C. van de Kamer and Dr. F. C. G. van de Veerdonk for their helpful encouragement and constructive suggestions, Dr. L. Boomgaart for checking and amending the English writing. Thanks are also due to Miss Ans de Groot and Miss Thera Verstappen for their technical assistance and to the staff of the photographic department for making the illustrations.     

Die Ontogenese der aminhaltigen Neuronensysteme im Gehirn von Rana temporaria

Zusammenfassung Vorkommen und Verteilung biogener Amine im Gehirn von Rana temporaria-Kaulquappen wurden fluoreszenzmikroskopisch untersucht. Catecholaminhaltige Perikaryen erscheinen ab Stadium 20 im Nucleus reticularis mesencephali, im Tuber cinereum und im Bulbus olfactorius, ab Stadium 22 in den Flügelplatten der Medulla oblongata und in der Area praeoptica. Ab Entwicklungsstufe 20 zeigen sich ventrolateral in Medulla oblongata und Mittelhirn, lateral vom Organon vasculosum hypothalami, im Bereich des medialen Vorderhirnbündels und im Striatum catecholaminhaltige Faseranschwellungen, ab Stadium 22 außerdem in der Eminentia mediana und dem Hypophysenzwischenlappen, in der Commissura transversa (bis zur Stufe 26), in der Commissura anterior (bis zur Stufe 26) und in der Pars ventrolateralis nuclei lateralis septi. Im Striatum ist von dieser Entwicklungsstufe an ein zweites Areal mit grün fluoreszierenden Varikositäten nachweisbar. Ab Stadium 26 finden sich auch in der Pars dorsolateralis des lateralen Septumkerns catecholaminhaltige Faseranschwellungen.Ab Entwicklungsstufe 22 sind 5-HT-haltige, gelb fluoreszierende Perikaryen im Nucleus raphes und in seiner Umgebung zu beobachten, gelb fluoreszierende Varikositäten im Nucleus interpeduncularis und zwischen medialem und lateralem Septumkern.

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Ontogeny of the amine-containing nerve cell systems in the brain of Rana temporaria

Summary The occurrence and distribution of biogenic amines in the brain of Rana temporaria tadpoles have been investigated with the fluorescence-microscope. From the embryonic developmental stage 20 onwards catecholamine-containing cell bodies are shown to be present in the nucleus reticularis mesencephali, the tuber cinereum and the olfactory bulb, and from stage 22 onwards also within the dorsolateral areas of the medulla oblongata and within the preoptic area. Catecholamine-containing enlargements of nerve fibres occur in the ventrolateral parts of the medulla oblongata and the midbrain, in an area lateral to the hypothalamic organon vasculosum, within the region of the medial forebrain bundle and within the striatum, in all stages following stage 20. These enlargements also occur in the median eminence and the pars intermedia of the hypophysis, in the commissura transversa (up to stage 26), in the commissura anterior (also up to stage 26) and in the pars ventrolateralis nuclei lateralis septi in all stages after 22. From the same stage onwards a second area of green fluorescent varicosities can be demonstrated within the striatum. After stage 26 catecholamine-containing enlargements of nerve fibres additionally are to be found in the dorsolateral part of the lateral septal nucleus.After appearing at stage 22 5-HT-containing, yellow fluorescent perikarya are to be observed within the nucleus raphes and its neighbourhood, and yellow fluorescent varicosities in the interpeduncular nucleus and in an area between the medial and the lateral septal nucleus.

Herrn Professor Dr. med. W. Bargmann zum 65. Geburtstag gewidmet.     

Identification of adenohypophysiotropic neurohormone producing neurosecretory cells in Rana temporaria

Summary By means of electron microscopy, in the median eminence of Rana temporaria, the terminal arborizations of axons of six different types of neurosecretory cells, located in the pars ventralis of the tuber cinereum, were identified. In addition, phenomena connected with the release of neurosecretory material from the axon terminals of these neurosecretory cells into the blood capillaries of the median eminence are described.Preliminary results suggested the existence, in the median eminence, of additional different neurosecretory axon types which could also belong to corresponding neurosecretory cell types probably located in the apical part of the pars ventralis of the tuber cinereum. Moreover, in the external region of the normal median eminence, separate monoaminergic nerve fibres were tentatively identified. Arguments are adduced which plead (1) against the assumption that the ependyma or the pituicytes of the median eminence could produce adenohypophysiotropic hormones; (2) against the inference that the ependymal cells of the median eminence might be involved in the transport of adenohypophysiotropic hormones from the cerebrospinal fluid into the blood capillaries of the median eminence.     

Seasonal changes of the ultrastructure of the pars tuberalis of the hypophysis of Rana temporaria

Summary In the pars tuberalis of the hypophysis of Rana temporaria, which shows the ultrastructural characteristics of a polypeptide hormone secreting endocrine gland, seasonal changes of the ultrastructure are described. In accordance with the literature, these seasonal changes of ultrastructure are interpreted as the morphological expression of seasonal changes of endocrine activity of the pars tuberalis.     

Electron microscopic immunocytochemical demonstration of separate vasotocinergic and mesotocinergic nerve fibres in the median eminence of the frog hypophysis

Summary With the use of the unlabelled antibody peroxidase-antiperoxidase complex (PAP) technique at the electron microscopic level, it was shown that both the internal and the external regions of the median eminence of the frog contain separate vasotocinergic and mesotocinergic nerve fibres. This observation confirms the results of previous immunocytochemical studies at the light microscopic level. The mean size of the neurohypophysial hormone-containing granules in the external region of the median eminence is significantly smaller than that of the neurohypophysial hormone-containing granules in the internal region of the median eminence. No significant difference could be found between the mean granule size of the vasotocinergic and mesotocinergic fibres of either the internal or the external region of the frog median eminence.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

Immunoreactive Tamm-Horsfall protein in the kidney and skin of the frog Rana temporaria

Summary Tamm-Horsfall protein (THP) is the main protein in normal human urine, and is found in the thick limb of the Loop of Henle in human kidney, and in other mammalian species. The skin of the frog, Rana temporaria, has similar physiological properties to this mammalian kidney tissue. In the present study, an immunohistological method involving an antibody to human THP was used to investigate the distribution of this distinctive protein in frog kidney and skin, and to compare its distribution with that found in the kidney tubules of rat and rabbit. THP-positive material was detected in the distal renal tubules and nephric duct of frogs, and was also located in the superificial epidermis of skin. It is suggested that its presence in amphibian skin is consistent with the hypothesis that THP is an important component of tissues that absorb sodium and chloride ions, but remain impermeable to water.     

Sound and vibration sensitivity of VIIIth nerve fibers in the grassfrog, Rana temporaria

We have studied the sound and vibration sensitivity of 164 amphibian papilla fibers in the VIIIth nerve of the grassfrog, Rana temporaria. The VIIIth nerve was exposed using a dorsal approach. The frogs were placed in a natural sitting posture and stimulated by free-field sound. Furthermore, the animals were stimulated with dorso-ventral vibrations, and the sound-induced vertical vibrations in the setup could be canceled by emitting vibrations in antiphase from the vibration exciter. All low-frequency fibers responded to both sound and vibration with sound thresholds from 23 dB SPL and vibration thresholds from 0.02 cm/s². The sound and vibration sensitivity was compared for each fiber using the offset between the rate-level curves for sound and vibration stimulation as a measure of relative vibration sensitivity. When measured in this way relative vibration sensitivity decreases with frequency from 42 dB at 100 Hz to 25 dB at 400 Hz. Since sound thresholds decrease from 72 dB SPL at 100 Hz to 50 dB SPL at 400 Hz the decrease in relative vibration sensitivity reflects an increase in sound sensitivity with frequency, probably due to enhanced tympanic sensitivity at higher frequencies. In contrast, absolute vibration sensitivity is constant in most of the frequency range studied. Only small effects result from the cancellation of sound-induced vibrations. The reason for this probably is that the maximal induced vibrations in the present setup are 6–10 dB below the fibers' vibration threshold at the threshold for sound. However, these results are only valid for the present physical configuration of the setup and the high vibration-sensitivities of the fibers warrant caution whenever the auditory fibers are stimulated with free-field sound. Thus, the experiments suggest that the low-frequency sound sensitivity is not caused by sound-induced vertical vibrations. Instead, the low-frequency sound sensitivity is either tympanic or mediated through bone conduction or sound-induced pulsations of the lungs.Abbreviations AP amphibian papilla - BF best frequency - PST peristimulus time     

Cyclical mitotic activity in the adeno-hypophysis of Rana temporaria

Summary In the pars distalis, the pars intermedia and the pars tuberalis of the hypophysis of Rana temporaria, an annual mitotic cycle is described.In the pars distalis there exists a gradient of mitotic activity, corresponding to a gradient of distribution of the PAS positive cells.In the pars distalis, the rate of cell degeneration and of cell renewal of the PAS positive cells seems to be faster than of the orange G positive cells.The replacement of degenerated PAS positive cells of the pars distalis seems to occur mainly during the months immediately following on the period of maximal activity of the PAS positive cells.     

Arterial supply of the choriocapillaris of anuran amphibians (Rana temporaria,Rana esculenta)

Summary The pattern of the vascular supply to the choroid of the frog eye was studied in toto with the use of the injection-replication-SEM technique. The choroid of anuran amphibians is composed mainly of the choriocapillaris. In both species studied (Rana temporaria, Rana esculenta), an independent arterial supply to the choriocapillaris supplemented that from the ciliary arteries. This additional vascular route arises from the optic artery, a separate branch of the arteria infundibularis superficialis. The optic artery, accompanied by its vein within the vascular sheath of the optic nerve, joins the rich arterial capillary network of the choriocapillaris and supplies the posterior pole of the ocular bulb. The superficial capillary network displays a dense collar around the entrance of the optic nerve into the eye and is composed of a circular meshwork of small capillaries, several layers deep. More peripherally, however, it becomes single layered. This capillary network, as a whole, establishes numerous connections with the adjacent choriocapillaris at the posterior pole of the ocular bulb. In anuran amphibians the complex arrangement of both arterial systems supporting the choriocapillaris may be regarded as a more complete equivalent of the short posterior ciliary arteries of mammals.     

Weight loss,reproductive output,and the cost of reproduction in the common frog,Rana temporaria

Summary The consequences of reproduction for body weight, growth and survival were studied in a Swiss population of the explosive breeder, Rana temporaria. Males and females continuously loss weight in the range of 0.5% of total body weight per day from the breeding migration throughout May. Females also lost about 33% (1983) and 29% (1984) due to spawning. In addition to this significant year-to-year variation, there was also considerable individual variation in reproductive output. Skeletochronological techniques indicated that breeding male or female frogs experienced a growth reduction of several millimeters relative to non-breeding frogs of the same body size. There was no relationship between an individual female's reproductive output in consecutive years or with her subsequent growth or survival. It was concluded that weight loss is caused by a seasonally elevated metabolism in combination with a lack of feeding and represents a basic energetic cost of reproduction, resulting in lowered growth. Individual variation in relative reproductive output is mostly environmentally induced and is not an expression of different reproductive strategies. This may explain the lack of trade-offs that are predicted by the cost-of-reproduction-hypothesis.     

Electron-microscopic investigation of the innervation of the pars distalis of the hypophysis in adult Rana temporaria L.

Summary In the pars distalis of the hypophysis of adult Rana temporaria, three types of nerve-fiber profiles were found at two distinct sites, in both lateral parts of the bordering regions of the anterior lobe with the intermediate lobe of the hypophysis. The first type of nerve-fiber profile consists of bundles of very fine axonal elements (diameter: <0.7 m). The second type is formed by larger nerve fibers (diameter up to 4 m) containing a few neurosecretory granules of approximately 100 nm. The third type of nervefiber profile resembles the second type but these nerve fibers make synaptoid contacts on at least two different types of glandular cells. The possible functional significance of these nerve fibers in the pars distalis is discussed.No nerve fibers were found (1) in the central part of the bordering region of the pars distalis with the intermediate lobe, (2) at the bordering region with the median eminence and (3) with the neurohypophysial stalk, and (4) in all other parts of the pars distalis.     

Amiloride-sensitive Na+/H+ exchange stimulated by cellular acidification or shrinkage in red blood cells of the frog,Rana temporaria

Simultaneous net uptake of Na⁺ and net extrusion of H⁺, both inhibited by amiloride, could be stimulated in red blood cells of the frog, Rana temporaria, either by intracellular acidification or cellular shrinkage. Net transports of Na⁺ and H⁺ were transient, dying out after 10–20 min (20°C) when stimulated by intracellular acidification but developing more slowly and proceeding for more than 60 min (20°C) when stimulated by cellular shrinkage. Evidence is presented suggesting a coupling between the transports of Na⁺ and H⁺ with an exchange ratio of 1:1 Na⁺/H⁺ exchange, stimulated by intracellular acidification, was able to readjust intracellular pH also when operating in parallel to a fully working anion exchanger in CO₂/HCO ₃ ^- -buffered media. Inhibition of anion exchange resulted in reduced cellular net uptake of Na⁺.Abbreviations DIDS 4,4-diisothiocyanatostilbene-2,2-disulphonate - DMSO dimethylsulphoxide - IU international unit - pH _e extracellular pH - pH _i intracellular pH - RBC red blood cell     

Scanning electron microscopy of the wall of the third ventricle in the brain of Rana temporaria. Part IV.

Summary The surface specializations of the wall of the third cerebral ventricle of Rana temporaria were investigated with the scanning electron microscope. These specializations can be divided into three types: cilia, large bulbous protrusions, and microvillus-like protrusions.Most parts of the ventricular surface are densely ciliated. In contrast, other regions are either scantily ciliated or devoid of cilia. Four areas of the ventricular surface are studded with numerous large bulbous protrusions. These large protrusions can be divided into two types: One type consists of intraventricular end bulbs of dendrites of secretory neurons. The other type is represented by large cytoplasmic extensions of ependymal cells.In the third ventricle of Rana, microvillus-like surface specializations of ependymal cells are ubiquitous structures. Generally, filiform protrusions of varying length are the predominant type. The microvillus-like specializations are transient structures, the number of which varies according to different physiological states of the ependymal cells.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

Patterns of cellular proliferation and migration in the developing tectum mesencephali of the frog Rana temporaria and the salamander Pleurodeles waltl

The development of the tectum mesencephali was studied in the frog Rana temporaria and the salamander Pleurodeles waltl by means of nuclear staining and by labeling of cells with bromodeoxyuridine (BrdU). The general spatial and temporal pattern of cell proliferation and cell migration is the same in both species, despite drastic differences in overall tectal morphology. However, the salamander species differs from the frog species by (1) a generally lower cell proliferation rate, (2) a reduction in the activity of the lateral proliferation zone, and (3) a reduction in the formation of superficial cellular layers. Because point (3) affects processes that occur late in ontogeny, our experiments provide evidence that the simple morphology of the tectum of Pleurodeles waltl, compared with the multilayered tectum of Rana, is a consequence of a paedomorphic alteration of the ancestral developmental pattern of the amphibian tectum.     

Cell junctions with funnels in the olfactory mucosa of frogs (Rana temporaria L.)

Summary A characteristic structure in the apical junctional belt of the olfactory epithelium in Rana temporaria is visible in freeze-fracture preparations. This structure is described as a funnel with channel across the junctional belt. It is supposed to represent a possible way for discarding used molecules after stimulation, and to allow the stimulation of free nerve endings in the depth of olfactory epithelia.I wish to thank Prof. C.F. Bardele and Mr. H. Schoppmann for their kind support and technical help