The distribution and colocalization of neuropeptides and catecholamines in nerves supplying the gall bladder of the toad,Bufo marinus

The indirect immunofluorescence technique was used to determine the distribution of peptide-containing axons in the gall bladder of the cane toad, Bufo marinus. In addition, the adrenergic innervation of the gall bladder was examined by use of immunoreactivity to the catecholamine-synthesizing enzyme, tyrosine hydroxylase, and glyoxylic acid-induced fluorescence. On the basis of peptide coexistence, two intrinsic populations of neurones and their projecting fibres could be distinguished substance P neurones and vasoactive intestine peptide neurones. Neither of these two types of neurones contained any other colocalized neuropeptides. Four populations of nerve fibres arising from cell bodies outside the gall bladder were identified: nerves containing colocalized galanin, somatostatin and vasoactive intestinal peptide; nerves containing colocalized calcitonin gene-related peptide and substance P; adrenergic nerves containing neuropeptide Y; and nerves containing only adrenaline.     

Visual performance of the toad (Bufo bufo) at low light levels: retinal ganglion cell responses and prey-catching accuracy

The accuracy of toad snapping towards moving worm dummies under various levels of dim illumination (from absolute threshold to moonlight) was videorecorded and related to spike responses of retinal ganglion cells exposed to equivalent stimuli. Some toads (at ca. 16 °C) successfully snapped at dummies that produced only one photoisomerization per 50 rods per second in the retina, in good agreement with thresholds of sensitive retinal ganglion cells. One factor underlying such high sensitivity is extensive temporal summation by the ganglion cells. This, however, is inevitably accompanied by very long response latencies (around 3 s near threshold), whereby the information reaching the brain shows the dummy in a position where it was several seconds earlier. Indeed, as the light was dimmed, snaps were displaced successively further to the rear of the dummy, finally missing it. The results in weak but clearly supra-threshold illumination indicate that snaps were aimed at the advancing head as seen by the brain, but landed further backwards in proportion to the retinal latency. Near absolute threshold, however, accuracy was too good, suggesting that the animal had recourse to a neural representation of the regularly moving dummies to correct for the slowness of vision.     

Effects of NEM on Voltage-activated Chloride Conductance in Toad Skin

The regulation of the voltage-activated chloride current conductance (G _Cl ) in toad skin was investigated by the use of the SH reagents N-ethylmaleimide (NEM) and p-chloro-mercuricbenzenesulfonic acid PCMBS. This anion pathway is controlled by a voltage-sensitive gating regulator. Mucosal application of NEM decreased the voltage-activation in a time and concentration dependent manner, half-maximal inhibition being exerted at a concentration of 30 μm within 20 min. At concentrations higher than 100 μm, the voltage-activated G _Cl was near-completely and irreversibly inhibited in less than 10 min. Resting, deactivated conductance was essentially unaffected. NEM had no effect on active sodium transport (measured as I _sc ) under conditions, which fully dissipated the voltage-activated G _Cl . After complete inhibition of the voltage-activated G _Cl with NEM, chloride conductance could still be stimulated by CPT-cAMP as in control tissues. Under these conditions, NEM at concentrations above 1 mm decreased G _Cl reversibly. Mucosal application of PCMBS at 500 μm inhibited the activated conductance by 35%, which was slightly reversible. Inhibition of voltage-activated G _Cl , which was observed after mucosal addition of the membrane-impermeable NEM analogue, eosin-5-maleimide, was completely reversible after washout. This suggests that the binding site for the maleimide is not accessible from the external face of the apical membrane. Brief application of NEM at lower concentrations (1–3 min, ≤100 μm) led to partial inhibition of G _Cl , followed by occasionally complete recovery upon washout of NEM. Recovery of voltage-activated G _Cl was progressively attenuated and eventually disappeared after subsequent brief applications of NEM. This could reflect recruitment of permeation/control sites from a finite pool. The data are discussed in the frame of a working model for the voltage-activated Cl⁻-pathway, that contains two principle components, i.e., an anion-selective permeation path which is controlled by regulatory protein(s). Received: 18 December 1996/Revised: 28 April 1997     

文昌鱼轮器哈氏窝匀浆对幼体蟾蜍睾丸发育的初步探讨

关于文昌鱼生殖活动的调节,Tjoa和Welsch根据电镜的观察,推测文昌鱼头部轮器上的哈氏窝(Hatschek’s pit)可能与脊椎动物的脑下垂体同源。最近,张致一等用荧光免疫法研究发现轮器上的哈氏窝上皮细胞有与LH抗血清产生沉淀反应的分泌颗粒,从而进一步证实了Tjoa等人的看法。我们在光学显微镜下也观察到哈氏窝上皮细胞内分泌颗粒的数量,在性腺发育的不同阶段有明显的区别(待发表的资料),提示哈氏窝上皮细胞与性腺发     

Fine structural and cytochemical study of the innervation of smooth muscle in an amphibian (Bufo marinus) lung before and after denervation

Summary The innervation of the toad (Bufo marinus) lung was studied with transmission electron microscopy and fluorescence techniques, both before and after 12 or 20 days close vagosympathetic denervation. Four cytologically distinct types of neuronal processes were recognised, in relation to the visceral muscles of the lung. These were described as cholinergic, adrenergic, nonadrenergic/non-cholinergic (NANC) and sensory on the basis of the characteristics of their vesicular content and cytochemical reactions. An apparent efferent innervation of visceral smooth muscle was achieved by NANC (50%), cholinergic (25%) and adrenergic (25%) fibres. A few sensory fibres were also present. After denervation only NANC fibres persisted, showing that the cell bodies of these fibres were intrapulmonary. The vascular smooth muscle was supplied by cholinergic, adrenergic and sensory fibres. In the walls of the proximal branches of the pulmonary artery were fibres containing large dense-cored vesicles. These profiles, which were associated with the vasa vasorum, were similar to neurosecretory fibres. After denervation all neural profiles associated with the vasculature had degenerated. The observations suggest that vagal vasodepressor effects in the toad lung are mediated indirectly through relaxation of visceral muscle strands which in their contracted state compress vascular channels.The authors would like to thank Dr. J.R. McLean for technical advice on fluorescence microscopy. This work was supported by a grant from the Australian Research Grants Committee     

Innervation and cytochemistry of the neuroepithelial bodies in the ciliated epithelium of the toad lung (Bufo marinus)

Summary Neuroepithelial bodies (NEB) were identified in the lung of Bufo marinus. The characteristics of the cells and their innervation were studied with electron and fluorescence microscopy before and after close vagosympathetic denervation. The bodies consist of low columnar cells which rest on the epithelial basal lamina. The majority of the cells do not reach the lumen of the lung (basal cells); the few which do (apical cells) are bordered by microvilli and possess a single cilium. The neuroepithelial cell cytoplasm contains a variety of organelles the most characteristic of which are dense cored vesicles. Microspectrofluorometry and electron microscopic cytochemistry indicate significant quantities of 5-hydroxytryptamine in these cells. The neuroepithelial bodies could be divided into three groups on the basis of their innervation: 1) About 60% of the NEBs are innervated solely by nerve fibres containing agranular vesicles which form reciprocal synapses; 2) about 20% are innervated solely by adrenergic nerve fibres which form distinct synaptic contacts; and 3) the remaining 20% are innervated by both types of nerve fibres. It is proposed that the NEBs are receptors monitoring intrapulmonary P_{CO 2} and so leading to modulation of activity in afferent nerve fibres (type containing agranular vesicles). The presence of NEBs solely with an adrenergic (efferent) innervation poses a problem with this interpretation.     

The vascularization of the neural stalk and the pars nervosa of the hypophysis in the toad,Bufo bufo (L.) (amphibia,anura)

The angioarchitecture of the neural stalk and the encephaloposthypophysial portal system of the hypophysis of the toad, Bufo bufo (L.), was studied using three different methods. The neural stalk is mainly supplied by branches of the arteria infundibularis superficialis which form a widemeshed vascular network. Dorsally this network continues into the plexus of the pars nervosa. The vascularization of the pars nervosa is made up of the encephalo-posthypophysial portal system. This portal system consists of a hypothalamic branch (=portion), a mesencephalic and a mesencephalicbulbar branch (=portion). The hypothalamic branch was found to drain the pars ventralis of the tuber cinereum as well as more dorsal regions of the diencephalon. The mesencephalic-bulbar trunk enters the hypothalamic branch. The resulting common stem of the encephalo-posthypophysial portal vein the curves around the retroinfundibular communicating artery, crosses its ventral side and runs caudally. The secondary capillary plexus of the pars nervosa is characterized by well defined capillary plexus of the pars nervosa is characterized by well defined capillary networks which are located at the periphery of the parenchyma of the pars nervosa, thus forming a rostral, dorsal and ventro-caudal net. The central region of the parenchyma of the pars nervosa is supplied only by main branches of the encephalo-postpophysial portal vein. The venous drainage of the pars nervosa is via the vena hypophysea transversa.     

Histochemistry of the juxtaglomerular apparatus in the toad Bufo bufo

Summary An investigation regarding the question of whether there exists a macula densa as part of the juxtaglomerular apparatus in the kidney of amphibians has been carried out. With the aid of a histochemical reaction for glucose-6-phosphate dehydrogenase activity, the presence of a macula densa zone as a specialized part of the distal tubule in the toad Bufo bufo was demonstrated. The functional significance of the high glucose-6-phosphate dehydrogenase activity in the macula densa cells is discussed.     

Capacitance,short-circuit current and osmotic water flow across different regions of the isolated toad skin

Summary The amphibian antidiuretic hormone, arginine vasotocin, stimulated osmotic water flow across isolated skin from the pelvic but not the pectoral skin of the toad, Bufo woodhouseii. Changes in the apical membrane capacitance were not observed for either region of the skin following treatment with arginine vasotocin when there was an osmotic gradient across the tissue. In the absence of an osmotic pressure gradient, the apical membrane capacitance of the pelvic skin increased from 2.8±0.5 to 3.3±0.6 F · cm^-2 after treatment with 5 · 10^-8 M arginine vasotocin. Under these conditions, apical membrane capacitance of the pectoral skin was 1.8±0.1 F · cm^-2 and did not change significantly after arginine vasotocin treatment. The amiloride-sensitive short-circuit current across the pelvic skin was stimulated by arginine vasotocin as was the density of channels in the apical membrane as determined by fluctuation analysis. Values for channel density in the pelvic skin also correlated with apical membrane capacitance and increased from 90 to 273 channels per m² of estimated membrane area following arginine vasotocin treatment. In the pectoral skin the stimulation of short-circuit current following arginine vasotocin treatment was small and an increase in channel density could not be demonstrated. The current through single Na⁺ channels in both regions of the skin did not different either before or after arginine vasotocin treatment.Abbreviations A amiloride - ADH antidiuretic hormone - AVT arginine vasotocin - C capacitance - C _a capacitance of apical membrane - f _c corner frequency - i single-channel current - osmotic water flow - IMP intramembrane particles - I _sc short-circuit current - amiloride-sensitive short-circuit current - M channel density - P _o probability of a channel being open - R channel receptor - R _a apical resistance - R _p paracellular resistance     

Evidence for plasticity in the frequency of skipped breeding opportunities in common toads

Breeding is limited by energetic or environmental constraints and long-lived species sometimes skip breeding opportunities. Environmental conditions may vary considerably across the geographic and elevational range of a species and species that can respond through variation in life history strategies are likely to maintain populations at the extremes of their ranges. The decision to skip breeding enables animals to adjust life history to circumstances, and plasticity in behavior allows implementation of adjustments. Elevational patterns suggest that breeding may be limited physiologically at high elevations (e.g., greater probability of skipped breeding; resources and environmental conditions more variable) in contrast to low elevations (probability of skipping breeding lower; resources and environmental conditions more predictable). We estimated the probabilities of survival and skipped breeding in a high-elevation population of common toads and compared estimates to existing data for common toads at low elevations, and to another toad species inhabiting a similar high elevation environment. Female common toads at high elevations tend to have high probabilities of skipping breeding and survival relative to data for common toads at low elevations, and appear to use a similar strategy of skipping breeding in response to similar environmental constraints as other toads at high elevations. We provide evidence of variability in this aspect of life history for common toads. Understanding variation in life history within widely distributed species is critical. Knowing that certain life history strategies are employed on a continuum informs conservation efforts, especially as impacts of climate change are likely to be different depending on elevation.