Angiotensin converting enzyme in the toad Bufo arenarum

Angiotensin converting enzyme activity (ACEA) was determined in serum, kidney, whole skin and isolated epithelia homogenates of the South American toad Bufo arenarum. ACEA was present in the tissues and serum of the toad. The activity was higher in the kidney, as compared to that of the whole skin or isolated epithelium. Captopril, teprotide and EDTA, caused a significant decrease in the ACEA. Possible physiological roles for the presence of ACEA in the toad are discussed.     

Membrane currents in the oocyte of the toad Bufo arenarum

The amphibian oocyte cell model is widely used for heterologous expression of ionic channels and receptors. Little is known, however, about the physiology of oocyte cell models other than Xenopus laevis. In this study, the two-electrode voltage clamp technique was used to assess the most common electrical patterns of oocytes of the South American toad Bufo arenarum. Basal membrane resistance, resting potential, and ionic currents were determined in this cell model. The oocyte transmembrane resistance was 0.35 M(Omega), and the resting potential in normal saline was about -33 mV with a range between -20 mV and -50 mV. This is, to our knowledge, the first attempt to begin an understanding of the ion transport mechanisms of Bufo arenarum oocytes. This cell model may provide a viable alternative to the expression of ion channels, in particular those endogenously observed in Xenopus laevis oocytes.     

Antidiuretic responses to osmotic cutaneous stimulation in the toad,Bufo arenarum

Summary Osmotic stimulation of the skin of the toadBufo arenarum with isotonic (115 mM) or hypertonic (400 mM) NaCl solutions produced a marked and reversible antidiuresis within 5 min. No changes in plasma osmolarity were detected in the course of this response.Hypophysectomized animals exhibited a lower and delayed antidiuresis when exposed to a hypertonic environment (400 mM NaCl). This antidiuretic response was drastically reduced in normal toads after ten consecutive days of administration of the sympatoplexic guanethidine.The existence of a feed-forward control of urine production initiated by cutaneous osmotic sensors and involving an adrenergic component is proposed.     

Vascularization of the pars intermedia of the hypophysis in the toad,Bufo bufo (L.) (Amphibia,Anura)

The vascularization of the pars intermedia of the hypophysis of the toad, Bufo bufo (L.) was studied by injection of a mixture of India-ink and gelatine into the circulatory system of the head via the arteria carotis communis. Further methyl-methacrylate corrosion casts of brains were made and the hypophysial region of the corrosion casts was examined with the scanning electron microscope. The results showed that the vascularization of the pars intermedia of the toad hypophysis consists of a single-layered vascular network, which is located on the ventral surface of the pars intermedia. The network is formed by capillaries, which primarily run caudally in a fan-like manner and which show only a few cross-connections. In the rostral region of the pars intermedia this network lies rather superficially, while in the caudal region it slightly penetrates the parenchyma. The vascular network originates from vessels of the neural stalk and from wide capillaries of the rostro-ventral region of the neurointermediate junction. The venous drainage of the pars intermedia is exerted by veins, which leave the caudal region and drain into the veins leaving the venous pole of the pars distalis. The flat, wide meshed vascular net on the ventral side of the pars intermedia, demonstrated in this study, fits into the concept that the pars intermedia of the anuran hypophysis is under the control of nerve fibers coming from the hypothalamus.     

The vitelline envelope-to-fertilization envelope transformation in the toad Bufo arenarum.

An investigation of some changes associated with the transformation of the vitelline envelope into the fertilization envelope in the egg of the toad Bufo arenarum is reported here. In most of the experiments described, the parameter used to demonstrate these changes was the stability of structural integrity of isolated envelopes when submitted to different agents and conditions. The envelopes used for this purpose exhibited a high degree of purity and remained apparently unaltered by the isolation procedure. As a quantitative method to ascertain their solubility rate, the release of uv-absorbing materials into solution was determined. Compared to the vitelline envelope, the fertilization envelope has proven to be less soluble in water, more stable in the presence of the chaotropic ion thiocyanate, and less susceptible to digestion in the presence of sperm lysin, trypsin, and pronase. In Bufo arenarum, as in other species, the vitelline envelope appears to be composed of glycoproteins. In contrast to previous results, however, disulfide bonds do not seem to be involved in their structural integrity. Thus, experiments carried out with isolated envelopes as well as with envelopes in situ have demonstrated a lack of effect of disulfide bond breaking agents on envelope stability. Evidence is presented suggesting that the solubility of envelopes in mercaptan solutions, as reported by other laboratories, is likely to be the expression of artifactual results.     

Exocytosis of cortical granules from activated oocytes of the toad,Bufo arenarum

Summary Observation of the cortical region of oocytes of Bufo arenarum by transmission electron microscopy reveals modifications on their surface and in the contents of the cortical granules (CG) during activation. In non-activated oocytes only amorphous cortical granules (ACG) can be observed. Activated oocytes display ACG, intermediate cortical granules containing both amorphous and membranous material (ICG), and a third type containing only membranous material (MCG). During exocytosis, CG release their contents into the perivitelline space, where the amorphous and membranous materials are found. The three types of CG found during oocyte activation suggest transformation of ACG to MCG and indicate that the different components of the cortical granules, when released into the perivitelline space, might play different roles in prevention of polyspermy.Members of the Scientific Research Career of CONICET, R. Argentina.     

Prolactin binding sites in the brain and kidneys of the toad,Bufo arenarum Hensel

Summary (¹²⁵I)-ovine prolactin (oPRL) binding was found in several brain areas of the toad,Bufo arenarum Hensel. The olfactory bulb, cerebral hemispheres, and both dorsal and ventral mesencephalic regions showed saturable, high affinity, (¹²⁵I)-oPRL binding, ranging between 5.6 to 29.9 fmol/mg protein, while the association constant (K _a) by Scatchard analysis was between 4.0 to 8.7×10⁹ M⁻¹. This binding was compared with the Scatchard plot of the kidney, which has been already described by other groups, and gave 41.7 fmol/mg protein andK _a 2.5×10⁹ M⁻¹. Liver showed no binding and in the cerebral hemispheres (¹²⁵I)-oPRL was not displaced by non-lactogenic hormones, indicating that binding was hormone and tissue specific.     

The ACTH of the pituitary pars intermedia]

The present investigation suggests that the intermediate lobe contains corticotrophic cells that are morphologically similar to those of the adenohypophysis. It can also be concluded that the ACTH molecules extracted from the intermediate lobe do not differ from those from the adenohypophysis. The immunocytochemical studies confirm the presence of ACTH in the intermediate lobes of all species studied so far, and stimulate the question of the mechanism of ACTH synthesis at this level. The interrelationships between different peptides of the intermediate lobe, including LPH, ACTH, CLIP and MSH, require further investigations. The intermediate lobe synthesizes and releases ACTH in vitro. This release can be stimulated by crude extracts of the median eminence or of cerebral cortex, and does not seem to be controlled by the CRF. Although ACTH of the intermediate lobe intervenes in vivo in the response to a neurogenic stress, one may not assume that the intermediate lobe alone plays an important role in the maintenance of the adrenal cortex, as the adrenals are similarly affected by hypophysectomy or pars distalis removal.