Location of acetylcholinesterase in axotomized ciliary ganglia

In the present note we have investigated the cytochemical localization of acetylcholinesterase (AChE) in the chick ciliary ganglion (CG) after post-ganglionic axotomy obtained by ablation of the eyeball. Preliminary results show at quite early stages after axotomy a remarkable reduction of cytoplasmic AChE, the residual one being localized in the rough endoplasmic reticulum. On the contrary synaptic areas, in particular those concerning the calyciform synapses, still show a marked AchE activity, similarly to what observed in physiological conditions. The decrease of cytoplasmic AChe in axotomized CG does suggest the possibility that such AChE undergoes to a topographical rearrangement moving towards the synaptic areas of ganglionic neurons.     

Patch-clamp study of isolated taste receptor cells of the frog

Summary Taste discs were dissected from the tongue ofR. ridibunda and their cells dissociated by a collagenase/low Ca/mechanical agitation protocol. The resulting cell suspension contained globular epithelial cells and, in smaller number, taste receptor cells. These were identified by staining properties and by their preserved apical process, the tip of which often remained attached to an epithelial (associated) cell. When the patch pipette contained 110mm KCl and the cells were superfused with NaCl Ringer's during whole-cell recording, the mean zero-current potential of 22 taste receptor cells was –65.2 mV and the slope resistance 150 to 750 M. Pulse-depolarization from a holding voltage of –80 mV activated a transient TTX-blockable inward Na current. Activation became noticeable at –25 mV and was half-maximal at –8 mV. Steady-state inactivation was half-maximal at –67 mV and complete at –50 mV. Peak Na current averaged –0.5 nA/cell. The Ca-ionophore A23187 shifted the activation and inactivation curve to more negative voltages. Similar shifts occurred when the pipette Ca was raised. External Ni (5mm) shifted the activation curve towards positive voltages by 10 mV. Pulse depolarization also activated outward K currents. Activation was slower than that of Na current and inactivation slower still. External TEA (7.5mm) and 4-aminopyridine (1mm) did not block, but 5mm Ba blocked the K currents. K-tail currents were seen on termination of depolarizing voltage pulses. A23187 shifted theI _K(V)-curve to more negative voltages. Action potentials were recorded when passing pulses of depolarizing outward current. Of the frog gustatory stimulants, 10mm Ca caused a reversible 5-to 10-mV depolarization in the current-clamp mode. Quinine (0.1mm, bitter) produced a reversible depolarization accompanied by a full block of Na current and, with slower time-course, a partial block of K currents. Cyclic AMP (5mm in the external solution or 0.5 m in the pipette) caused reversible depolarization (to –40 to –20 mV) due to partial blockage of K currents, but only if ATP was added to the pipette solution. Similar responses were elicited by stimulating the adenylate cyclase with forskolin. Blockage of cAMP-phosphodiesterase enhanced the response to cAMP. These results suggest that cAMP may be one of the cytosolic messengers in taste receptor cells. Replacement of ATP by AMP-PNP in the pipette abolished the depolarizing response to cAMP. Inclusion of ATP--S in the pipette caused slow depolarization to –40 to –20 mV, due to partial blockage of K currents. Subsequently, cAMP was without effect. The remaining K currents were blockable by Ba. These results suggest that cAMP initiates phosphorylation of one set of K channels to a nonconducting conformation.     

Olfactory thresholds in unilaterally bulbectomized rats

Five normal and five unilaterally bulbectomized rats were testedfor their intensity difference threshold, absolute thresholdand ability to acquire a two-odor discrimination. There wereno significant differences between groups for mean thresholdvalues or the rate at which the detection and discriminationtasks were acquired. These results do not support the notionproposed in earlier studiesthat central summation of birhinicallyinhaled odors increases sensitivity to threshold levels of odorants.     

P311 accelerates nerve regeneration of the axotomized facial nerve

In axotomized adult neurons, a process of axonal regrowth and re-establishment of the neuronal function has to be activated. Developmentally regulated factors may be reactivated during neuronal regeneration. Here we identify a gene, previously designated P311, that is up-regulated in the axotomized facial motoneurons. Ectopically expressed P311 localizes in the cytoplasm and the nucleus. Over-expression of P311 induces p21(WAF1/Cip1) expression, leading PC12 cells to differentiate and to have neuron-like morphologies. Adenovirus-mediated P311 gene transfer promotes neurite outgrowth of postnatal dorsal root ganglion neurons and embryonic hippocampal neurons in vitro. This effect is abolished by the activation of Rho kinase. P311 also facilitates nerve regeneration following facial nerve injury in vivo. Our data provide evidence that genes involved in the differentiation process contribute to the regeneration of injured mature neurons, and may provide a practical molecular target.     

Histochemical study of skeletal muscles of the frog, Rana temporaria

Histochemical profiles of muscles were identified based on staining for myosin ATPase activity. They reveal typical arrangement of muscular fibres with a zoned pattern. Tonic fibres have a unique histochemical profile and are mixed with the most oxydative fast fibres to form toxic zones. Muscles show fast profiles in thigh and tonic or mixed profiles in fore-arm.     

An ultrastructural study of the spleen of the ranid frog Rana perezi

The spleen of Rana perezi is encapsulated by connective tissue and shows by light microscopy two areas with no obvious border: the white pulp and the red pulp. The white pulp-lymphoid clusters are scattered throughout the organ and contain lymphocytes, reticular cells, and some plasma cells. The red pulp displays two different portions. The predominant region consists of reticular cells, lymphocytes, a variety of other leucocytes, and cells undergoing division. This area possibly performs a haemopoietic function. The smaller portion of the red pulp is characterized by reticular-phagocytic cells and may be haemocaretic in its function. Macrophages and pigmented cells occur in both white and red pulp. The organization of the spleen of R. perezi can be considered as a transitional or intermediate state between the primitive condition seen in certain fishes and amphibians and the more complex organ of ammiotes.     

An X-ray study of the pH property of frog sciatic nerve

Low-angle X-ray diffraction patterns have been recorded from frog sciatic nerve in pH solutions of 0.1–13.0. The normal X-ray pattern of frog sciatic nerve in Ringer's solution is maintained at pH 4.0–10.0. In acid pH, 2.5–4.0, and in alkaline pH, 10.0–11.0, the nerve myelin is in the partial swollen state. The partial swollen state and the normal state are reversible. Two physical states, the anomalous swollen state and the condensed state, at acid pH below 2.5 and the separated state at alkaline pH above 12.3 have been identified. These three physical states, the anomalous swollen state, the condensed state and the separated state, are reversible with each other on changing the pH solution but the normal state cannot be regained.     

Cytophotometric study of deoxyribonucleic acid in mesometral cells of experimental deciduoma in unilaterally pregnant rats]

Ploidy level of mesometral cells in experimentaly induced deciduomata is determinated by DNA-Feulgen cytophotometric measures. This tissue shown the first evolutive stages leading to polyplo?dy (mono-binucleated cells) it was unable to reach a plo?dy level higher than 4 n.     

Fat body of the frog Rana esculenta: An ultrastructural study

In the frog, the fat body is the largest body lipid deposit and is associated with the gonad. The aim of the present work was to investigate the fine structure of the fat body at different periods of the annual cycle and during prolonged starvation. Results indicate that fat body cells of Rana esculenta caught in autumn and after winter hibernation resemble mammalian adipocytes of white adipose tissue and contain markers of adipose tissue, such as S-100 protein and lipoproteinlipase. However, unlike mammalian adipocytes, fat body adipocytes consistently show small lipid droplets associated with their single, large lipid deposits, a lack of a definite external lamina, and the presence of cellular prolongations and spicula at their surfaces. Transmission and scanning electron microscopy in association with lanthanum tracer experiments suggest that in fat body adipocytes a vesicular-tubular system connects the cytoplasm and the interstitial space. In June (i.e., during the reproductive period), fat body adipocytes appear to have lost much of their lipid deposit and adjacent adipocytes show interdigitation of their plasma membranes and prominent Golgi complexes. In starved frogs, fat body cells can be almost devoid of lipid and in regression to a near-mesenchymal state. Nevertheless, these fat bodies still contain lipoproteinlipase activity (≈ 45% of that found in lipid-filled ones), indicating persistent adipose differentiation of the cells therein. Results presented here show that the R. esculenta fat body is an adipose organ undergoing reversible extreme changes in adipocyte fat content, which are associated with definite ultrastructural features. The fat body represents a suitable model for studying adipose tissue under different and extreme physiological conditions. © 1996 Wiley-Liss, Inc.     

Kinetic study of the interaction between frog epidermis tyrosinase and chloride

The effect of halide ions on frog epidermis tyrosinase has been characterized with the trypsin-activated enzyme. At pH 7, the order of inhibition is I- greater than Br- greater than Cl- greater than F-. Chloride, the most extensively studied halide, shows a competitive pattern with respect to the substrate, L-DOPA. Inhibition is strongly pH-dependent, with a pKa of 6.12 for the responsible protonatable group. Other kinetic constants are also calculated using a novel approach. The mechanism of interaction between chloride and the enzyme is discussed, and a model is proposed in which chloride interferes the tyrosinase activity by displacing a catalytically important ligand, probably a histidine residue of the side-chain, from the copper at the enzyme-active site.     

Comparative study of the conformational transitions of frog and rabbit phosphorylases B

1. Conformational motility of the purified muscle glycogen phosphorylase B from two species of vertebrates (rabbit and frog) was investigated by the Hydrogen-Exchange method and Infrared Spectometry. 2. The experimental results of the 1H-2H exchange were expressed in terms of the probability P of exposure to isotopic solvent of phosphorylase peptide groups and in terms of the corresponding changes in standard free energy delta Go. 3. The combined methods used didn't show considerable differences of the protein conformations in the physiological pH region but rabbit phosphorylase was only characterized by rather more compact structure in comparison with frog phosphorylase.     

Regeneration restores some of the altered electrical properties of axotomized bullfrog B-cells

In bullfrog B-type sympathetic neurones axon injury produces substantial changes in somal membrane properties. These include a shortening of action potential afterhyperpolarization (AHP) and an increase in action potential (AP) duration. In the present experiments we compared two injury situations: nerve crush, which was followed by regeneration, and nerve cut, after which regeneration to the original target was prevented, to investigate whether these electrophysiological changes were related to axon regeneration. Both crush and cut injuries produced a similar maximum decrease in AHP duration (to 33 and 30%) by 14 days after axotomy. After nerve crush, AHP duration recovered to within control values by 42 days, while after cut it remained depressed. AHP amplitude decreased to the same extent after nerve crush or cut (to 62 and 58%), but the rate of decrease was slower following crush when compared with cut, and following both types of injury it still remained depressed at 42 and 49 days. Changes in AP duration also took longer to occur following nerve crush, reaching maximal values at 35-42 days, at which time AHP duration had returned to within the normal range. The early reduction in AHP duration and its rapid recovery in regenerating neurones suggests that the current underlying this membrane property is regulated by events associated with axon outgrowth and peripheral reconnection. In contrast, changes in AHP amplitude and AP repolarization appeared to be independent of the occurrence of axon regeneration and remained abnormal at 49 days despite the recovery of AHP duration. These results imply that the electrophysiological changes seen in B-cells following injury are differentially regulated during subsequent regeneration.     

Stereologic estimates of glomerular volume in unilaterally nephrectomized rats

OBJECTIVE: To quantitatively evaluate the changes in glomeruli in unilaterally nephrectomized adult rats by unbiased stereologic techniques. STUDY DESIGN: Adult rats received unilateral nephrectomy (UN) or a sham operation. Four weeks later the remnant kidneys were fixed and sampled by the fractionator method. Consecutive sections were prepared and stained. Morphometric analyses were performed by a light microscope equipped with computer-assisted digitizer. RESULTS: The total glomerular volume increased by 51% (P < .01). The volume density of glomeruli did not change (5.58 +/- 0.22 x 10(-2) vs. 5.51 +/- 0.44 x 10(-2), P > .50). In contrast, the numerical densities of glomeruli decreased by 36% (P < .05). CONCLUSION: Glomerular enlargement, but not the increase in glomerular number contributed heavily to compensatory glomerular growth after UN in rats.