Pierre Deparis

Red blood cells and hemoglobin concentration in normal diploid and several types of polyploid salamanders

Abstract 1. 1. Different blood parameters were examined in both diploid and polyploid adult Pleurodeles newts. 2. 2. In 2 n, 3 n and 4 n, the hemoglobin content and hematocrit values were very close. 3. 3. The red blood cell counts decrease as the degree of polyploidy rises. 4. 4. Autotetraploid and pentaploid animals do not regulate their hemoglobin values and are slightly anaemic.

Erythrocyte enzyme activities in diploid and triploid salamanders (Pleurodeles waltlii) of both sexes

Studies of several enzyme activities and of some metabolites in erythrocytes of the newt Pleurodeles waltlii are described. The results indicate that differences exist between enzyme activities in males and females and in diploid and triploid animals. Females have higher erythrocyte enzyme activities than males. However, triploid animals have lower enzyme activities in their erythrocytes than diploid animals; this observation is more striking in females than in males.

Comparative plasma levels of androgens and 17β-estradiol in the diploid and triploid newt, Pleurodeles waltl

Seasonal changes in the plasma levels of androgens (testosterone plus dihydrotestosterone) and 17 beta-estradiol in diploid and triploid adult newts, Pleurodeles waltl were studied. In both male and female diploid individuals, large variations were reported with highest levels being found during breeding periods. In triploid newts seasonal variations were also found, similar to the diploid ones, but the plasma concentrations of the 17 beta-estradiol and androgens in triploid females and androgens in triploid males were lower throughout the year than those reported for the diploids. This difference is discussed in relation to the genetic sexual constitution.

Ontogeny of Amphibian Hemopoietic Cells

The developmental origin of lymphoid cells of vertebrates continues to be a fascinating, perplexing problem. Historically, the disputable issues have revolved around two fundamental aspects: the origin of the precursors of the various types of differentiated blood cells and the lineal relationships among these different cell types. There are special features of amphibians that make them suitable for clarifying the embryogenesis of hemopoietic cells. An array of microsurgical techniques from experimental embryology can be used to marked advantage. In this chapter, we will review the contributions made by investigators who have used the tools of experimental embryology to gain insight into the ontogeny of vertebrate blood cells.

Hemoglobin switching in the salamanderPleurodeles waltlii

SummaryIn order to study the cellular distribution of larval and adult hemoglobins during larval development ofPleurodeles waltlii a double specific immunofluorescent labelling technique was developed.Rabbit antibodies specific for larval and adult hemoglobin components were prepared and conjugated with tetramethyl-rhodamine isothiocyanate for the anti-larval antibodies and fluorescein isothiocyanate for the anti-adult hemoglobin antibodies.Both simultaneous and sequential staining with the two types of fluorescent antibodies indicated that larval and adult hemoglobins were never observed within the same erythrocyte during development. The results provide evidence that two distinct cell populations exist, one synthesizing exclusively larval hemoglobins which is progressively replaced by the other one synthesizing exclusively adult hemoglobins. It remains to be determined if these two populations arise from two distinct types of stem cells (adult and larval) or from the same stem cell type.

A genetic study of various enzyme polymorphisms in Pleurodeles waltlii (Urodele Amphibian). I. Lactate dehydrogenase-B and glucose-6-phosphate dehydrogenase

On starch gel electrophoresis of erythrocyte hemolysates of Pleurodeles waltlii (Urodele Amphibian), both lactate dehydrogenase-B (LDH-B) and glucose-6-phosphate dehydrogenase (G6PDH) show polymorphism that depends on a pair of autosomal codominant alleles, confirmed by analysis of gynogenesis progeny. Diploid gynogenesis results from fusion of the female pronucleus with the second polar body. The heterozygous state of a female for a given character is maintained in certain progeny when crossing-over occurs between the locus in question and the centromere. So the high proportion of heterozygotes (45.7% for LDH-B and 76% for G6PDH) indicates the high frequency of crossing-over and hence the large distance between each of the loci and the centromere.

Identification of the vitellogenin proteins in the newt Pleurodeles waltl (Urodele amphibian)

Vitellogenin derived from the blood of estrogen-treated Pleurodeles waltl was identified by immunochemical and electrophoretic analyses, using an antiserum against plasma vitellogenin isolated by dimethylformamide precipitation. Pleurodeles vitellogenin migrates as four bands on native PAGE, designated alpha-, beta-, gamma- and delta- VTG, with apparent mol. wts of 250,000, 270,000, 280,000 and 520,000 respectively. In the plasma, from estrogen-treated males like from ovariectomized estrogen-treated females, an additional band (mu-VTG) was found by native PAGE, never observed in estrogen-treated female plasma. It has a mol. wt of about 380,000 and shows complete immunological cross-reactivity with the vitellogenin antiserum. At least two polypeptides, termed VTG-I and VTG-II (mol. wt = 180,000 and 210,000) were identified by SDS-PAGE. Rocket immunoelectrophoresis displays three distinct precipitate lines indicating major immunological differences between the plasma vitellogenins.

Effects of thyroxine on the ontogeny of the vitellogenic response in Pleurodeles waltli of both sexes

SummaryIn the newt, Pleurodeles waltli, vitellogenin synthesis can be induced by oestrogen prior to metamorphosis and well before vitellogenin synthesis is normally observed in females. Vitellogenin indicubility has been shown to occur during normal development at stage 53 in both sexes. Immersion of young larvae in thyroxine at concentrations that did not result in early morphological metamorphosis led to a precocious acquisition of liver responsiveness to oestrogen. Treatment of larvae with thiourea greatly diminished the ability of oestradiol to induce vitellogenin synthesis, although it did not abolish the hormonal response.

Differential Protein Distribution Related to Dorsoventral Polarity in Pleurodeles waltl Cleaving Egg

The molecular basis for the initial specification of dorsoventral polarity in the Amphibian egg prior to the mid‐blastula transition still remains an open question. Regional differences in the protein pattern of Pleurodeles egg were investigated during early cleavage (8‐ and 512‐cell stages, prior to the mid‐blastula transition). Animal‐dorsal, animal‐ventral, vegetal‐dorsal and vegetal‐ventral quarters were separated and proteins were analyzed by 2D‐electrophoresis. The comparison of acidic protein patterns from dorsal and ventral quarters revealed differences between vegetal cells but no difference was detected between animal cells. One protein (p11, 30 kDa) was characterized in the dorsal side as early as the 8‐cell st. and two dorsal spots were detected at the 512‐cell st. (p11 and p5, 65 kDa). Similarly one protein (p7b, 46 kDa) appears to be ventral‐specific from the 8‐cell st. The p11 spot was shown to appear in ventral cells as a consequence of a dorsalizing LiCl‐treatment at the 32‐cell stage. Conversely, p11 disappeared from dorsal cells and p5 did not appear at 512‐cell stage after UV‐irradiation of the uncleaved egg, which results in the expression of the ventral‐specific protein p7b in the dorsal part of the egg. Therefore differential protein expression is already present at very early cleavage stages. Its significance needs further investigation.

Nuclear estrogen binding sites in the liver of the newt Pleurodeles waltl

Previous investigations of the liver estrogen-specific binding in the newt, Pleurodeles waltl, have identified in the cytosol fraction, only from normal males, a new middle-affinity estrogen-binding component (MEBC) displaying the properties of type II sites reported in various tissues of vertebrates. The present work demonstrates that MEBC sites are not unique to the male but are also present in the nuclei of female animals. However, comparative study between males and females of liver nuclear sites under various extraction conditions shows sex-linked differences in the subnuclear localization. The relationships between the association states of MEBC in the nuclear compartment according to sex and their presence or absence in the cytosol fraction are discussed.