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Objectives We report the pharmacological evaluation of a new series of 3-aminocoumarins differently substituted with hydroxyl groups, which have been synthesized because they include in their structures the tyrosine fragment (tyrosine-like compounds), with the aim of discovering structural features necessary for tyrosinase inhibitory activity. Methods The synthesized compounds 4 and 7–9 were evaluated in vitro as mushroom tyrosinase inhibitors. Key findings Two of the described compounds showed lower IC50 (concentration giving 50% inhibition of tyrosinase activity) than umbelliferone, used as a reference compound. Conclusions Compound 7 (IC50 = 53 µm) was the best tyrosinase inhibitor of this small series, having an IC50 value 10-fold lower than umbelliferone. Compound 7 (3-amino-7-hydroxycoumarin) had amino and hydroxyl groups precisely mimicking the same positions that both groups occupy on the tyrosine molecule.

We report the first case of cosegregation of two haemoglobins (Hbs): HbG-Philadelphia [α68(E17)Asn→Lys] and HbDuarte [β62(E6)Ala→Pro]. The proband is a young patient heterozygous also for β∘-thalassaemia. We detected exclusively two haemoglobin variants: HbDuarte and HbG-Philadelphia/Duarte. Functional study of the new double variant HbG-Philadelphia/Duarte exhibited an increase in oxygen affinity, with a slight decrease of cooperativity and Bohr effect. This functional behaviour is attributed to β62Ala→Pro instead of α68Asn→Lys substitution. Indeed, HbG-Philadelphia isolated in our laboratory from blood cells donor carrier for this variant is not affected by any functional modification, whereas purified Hb Duarte showed functional properties very similar to the double variant. NMR and MD simulation studies confirmed that the presence of Pro instead of Ala at the β62 position produces displacement of the E helix and modifications of the tertiary structure. The substitution α68(E17)Asn→Lys does not cause significant structural and dynamical modifications of the protein. A possible structure-based rational of substitution effects is suggested.

With the aim to find out structural features for the tyrosinase inhibitory activity, in the present communication we report the synthesis and pharmacological evaluation of a new series of phenylcoumarin derivatives with different number of hydroxyl or ether groups and bromo substituent in the scaffold. The synthesized compounds 5–12 were evaluated as mushroom tyrosinase inhibitors showing, two of them, lower IC50 than the umbelliferone. Compound 12 (IC50 = 215 μM) is the best tyrosinase inhibitor of this series.

Background Methylmalonic aciduria combined with homocystinuria (MMA–HC) is the biochemical trait of a metabolic disorder resulting from impaired conversion of dietary cobalamin (cbl, or vitamin B12) to its two metabolically active forms. Effects on urinary purine and pyrimidine levels have not been described for this condition. Methods Urine samples were collected from three patients with methylmalonic aciduria combined with homocystinuria and from 70 healthy subjects. Urinary purine and pyrimidine levels were quantitated by the use of LC/UV–Vis and LC/ESI/MS. Results Higher urine levels of pyrimidines were detected with both methods in patients compared to controls. Conclusion Methylmalonic aciduria with homocystinuria is due to deficiency of the enzyme, cobalamin reductase. The enzyme defect leads to altered hepatic metabolism, which appears to modify circulating pyrimidine levels.

Our study examines the functional and structural effects of amino acid substitution in the distal side of β-chains of human Hb Duarte (\documentclass[10pt]{article} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \pagestyle{empty} \oddsidemargin -1.0in \begin{document} \begin{equation*}{\alpha}_{2}{\beta}_{2}^{62{\mathrm{Ala}}{\rightarrow}{\mathrm{Pro}}}\end{equation*}\end{document}). We have compared the functional properties of the purified Hb Duarte with those of HbA, and through proton NMR and molecular dynamics simulations we have investigated their tertiary and quaternary structures. The variant exhibits an increased oxygen affinity with a normal Hill coefficient and Bohr effect. The abnormal function of Hb Duarte is attributed to the presence of a proline residue at the β62 position, since the functional properties of another Hb variant in the same position, Hb J-Europa (β62Ala→Asp), have been described as normal. Thereafter 1H-NMR studies have shown that the β62 Ala→Pro substitution causes structural modifications of the tertiary structure of the β globins, leaving the quaternary structure unaltered. These results have been confirmed by extensive all-atom molecular dynamics simulations. All these findings lead to the conclusion that the β62 Ala→Pro substitution produces a destabilization of the E-helix extending downward to the CD corner. Particularly, a cavity near the distal histidine of the β-chains, connecting the heme pocket to the solvent, is affected, altering the functional properties of the protein molecule.

We report the sixth occurrence of Hb Belfast [beta15(A12)Trp--Arg], a mild, unstable beta chain variant, in a large family wherein nine subjects were affected. DNA analysis showed a TUG--AGG mutation at codon 15 of the beta-globin gene, confirming a Trp--Arg amino acid substitution. The oxygen affinity of the isolated variant was increased. The clinical phenotype is silent or very mild, the only clinical finding being an intermittent moderate jaundice.

The occurrence of several abnormal hemoglobins (Hbs) in the population of the island of Sardinia has been documented [[1]]. Because of continuing screening programs for hemoglobinopathies among new...

Haemoglobin (Hb) J-Sardegna [α50(CE8)His → Asp] is a haemoglobin variant characteristic of subjects from the island of Sardinia. Here we report a study of the functional properties of both fetal and adult Hb J-Sardegna. The results indicate that adult Hb J-Sardegna displays an oxygen affinity that is higher than that of adult Hb only in the presence of 2,3-diphosphoglycerate (2,3-DPG). On the contrary, at 20 °C, the oxygen affinity of fetal Hb J-Sardegna is identical to that of normal fetal haemoglobin, both in the presence and in the absence of 2,3-DPG. A significant difference between these two systems (i.e. a higher oxygen affinity of fetal Hb J-Sardegna) shows up very clearly only when temperature is increased to 37 °C. Hence in fetal Hb, the main effect of the amino acid substitution is a decrease in the overall enthalpy change of oxygenation. The results outline the role of the α1-β1 interface in assessing the thermodynamics of oxygen binding. The functional properties of both adult and fetal Hb J-Sardegna have been interpreted at the structural level in light of the results obtained by a computational modelling approach performed in comparison with HbA and Hb Aichi, a variant characterized by a different mutation [α50(CE8)His → Arg] at the same position.

We applied best fitting procedures to capillary electrophoresis (CE) mobility values, measured at varying acidic pH, of a set of 21 peptides with a molecular mass ranging from about 350 to 1850 Da. This method allowed the contemporary measurements of C-terminus and carboxylic group of the side-chain of aspartic and glutamic acid dissociation constants and of peptide Stokes radius at different protonation stages. Stokes radius was related to peptide molecular mass M at the power of a fractional coefficient, and best correlation was found at pH 2.25, the fractional coefficient being equal to 0.68. This value is close to that proposed by R. E. Offord (Nature 1966, 211, 591-593), who suggested a proportionality between the polymer Stokes radius and M(2/3). The coefficient value decreases at higher pH, reaching a value of 0.58 at pH 4.25, corresponding to a mean peptide conformational transition towards more compact structures as a consequence of C-terminus dissociation. The measurement of the dissociation constants of each peptide allowed us to determine the percentage error on peptide charge predictions performed utilizing mean dissociation constants. Even for the charge, the best predictive performance is obtained at the most acidic edge of the range of the pH studied, mainly at pH 2.25. Conclusively, this study shows that the best performance of predictive models for peptide CE mobility is obtainable in the very acidic pH range (2.25-2.50) and in the absence of electroosmotic flow, and that a satisfactory predictive equation of peptide electrophoretic mobility (m2V(-1)s(-1) is given by mu = 85.4(Z/M(0.68))10(-8).

Tyrosinases, better termed polyphenoloxidases (EC 1.14.18.1; monophenol, o-diphenol: oxygen oxidoreductase), are copper-containing monooxygenases catalyzing the o-hydroxylation of monophenols to the corresponding catechols (monophenolase activity) and the oxidation of catechols to the corresponding o-quinones (diphenolase activity).

To evaluate serum purine metabolite concentrations in patients affected by fibromyalgia syndrome (FMS) and the relationships between their levels and FM clinical parameters.Serum purine levels were quantified using LC/UV-vis in 22 fibromyalgic females (according to the American College of Rheumatology classification criteria) and 22 healthy females.Significantly higher serum inosine, hypoxanthine and xanthine levels (p0.001) and significantly lower serum adenosine (p0.05) were detected in the FMS patients vs healthy controls. Our data show a negative correlation between adenosine and Fibromyalgia Impact Questionnaire (FIQ).Study results suggest that purines, in particular adenosine and inosine, may be involved in pain transmission in fibromyalgia.

During a screening program for the identification of β-thalassemia (β-thal) carriers in Sardinia, Italy, we identified two subjects with increased hemoglobin (Hb) levels and an abnormal Hb variant. The same variant was detected in a family member. DNA sequencing revealed a TGTTGG mutation at codon 93 of the β-globin gene. Structural analysis demonstrated that the cystine residue at position 93 of the β chain was substituted by tryptophan. Since this amino acid substitution had not yet been reported, we designated this variant Hb Santa Giusta Sardegna for the place of birth of the subjects. This amino acid substitution occurs at the tyrosine pocket of the β chain as well as at the α1β2/α2β1 contact of the quaternary structure of the molecule. The presence of this Hb in the hemolysate causes an increased oxygen affinity, a slightly reduced Bohr effect and a reduced heme-heme interaction (n(50), Hill's constant) in comparison with those of Hb A.

The effects of pH, organic phosphates (2,3-diphosphoglycerate), and temperature on the functional properties of both adult and fetal hemoglobin Sassari alpha (Asp-126-->His) have been studied. The functional properties of the adult variant are characterized by the following: (i) an oxygen affinity higher than that of normal HbA in all the experimental conditions used; (ii) a dramatic reduction of homotropic interactions (n50 very close to unity); and (iii) a significant decrease of the effect of 2,3-diphosphoglycerate, which is 35% lower than that observed on HbA. The fetal variant shows an increased oxygen affinity compared with normal HbF and an almost abolished heme-heme interaction. The molecular basis of these functional differences is discussed in terms of the possible role played by the substitution of alpha (Asp-26-->His) on the stability of the R state of the molecule due to a decreased interaction at the level of alpha 1 alpha 2 contact.

Coumarin derivatives (IV) and (VII) are synthesized by the Perkin reaction using dicyclohexylcarbodiimide as dehydrating agent, followed by hydrolysis of the methoxyl groups.

A series of novel bis-salicylaldehydes were synthesised and evaluated as tyrosinase inhibitors using a tyrosinase-dependent l -DOPA oxidation assay. The bis-salicylaldehydes exhibited greater inhibitory activity than salicylaldehyde. Our data suggests that these novel compounds may serve as a structural template for the design and development of novel tyrosinase inhibitors.

Using xenon atoms as a biomolecular probe raises the concern of whether they may influence in some way the molecular and electronic structure of the system under study. In this paper, the relevance of guest-host interactions in xenon complexes with paramagnetic myoglobins (Mbs) is thoroughly analyzed, and the issue about the use of xenon to detect and characterize voids within flexible biomolecules is critically discussed. A detailed 1 H NMR study useful for describing the hydrophobic cavities close to the active site of low-spin ferric myoglobins with respect to their interaction with the xenon atom is presented. The method is subsequently validated by the analysis of Xe-Mb with two different myoglobins, extracted from horse and pig. These myoglobins differ by 14 amino acids. One of these, Ile142 in horse Mb, is located in the proximal cavity, which is the main xenon binding site in horse Mb, and is replaced by Met142 in pig Mb. We demonstrated specific behaviors associated with the capacity of each of the two myoglobins to bind xenon and provided site-specific information on the host-guest interaction. Moreover, 1 H NMR measurements produce a picture of xenon-related local distortions of the protein, associated with a functionally relevant residue located right at the active site, the proximal hystidine E7(His93). According to the 1 H NMR data, xenon induces the tilt of the residue His93 relative to the heme plane and consequently causes an alteration of the magnetic axes. Similar conclusions are obtained both for pig cyano-myoglobin and for horse cyano-myoglobin, the structural deformation being in the former of minor entity.

In this work, the PEG-immobilization and the liquid phase synthesis of some coumarins derived from cardol are presented. Some preliminary results on their tyrosinase inhibitory activity are also included.

Tyramine, an important plant intermediate, was found to be a substrate for two proteins, a copper amine oxidase and a peroxidase from Euphorbia characias latex. The oxidation of tyramine took place by two different mechanisms: oxidative deamination to p-hydroxyphenylacetaldehyde by the amine oxidase and formation of di-tyramine by the peroxidase. The di-tyramine was further oxidized at the two amino groups by the amino oxidase, whereas p-hydroxyphenylacetaldehyde was transformed to di-p-hydroxyphenylacetaldehyde by the peroxidase. Data obtained in this study indicate a new interesting scenario in the metabolism of tyramine.

Birds during normal sustained flight must be able to dissipate more than 8 times as much heat as during rest in order not to be overheated. The experiments reported in this note on the hemoglobin systems from two different birds indicate the existence of a molecular mechanism by which hemoglobin is used simultaneously for oxygen transport and heat dissipation.

A novel hemoglobin variant was observed in pure sheep ( Ovis aries ) breeds of the island of Chios (Greece), Egypt and Hungary. This silent variant was identified by gel electrophoresis and RP-HPLC of dissociated globin chains. Two Arg for Lys substitutions were detected, by means of MALDI TOF electrospray mass spectrometric analysis for the intact globins, at positions β66(E10) and β144(HC1) of a globin chain having the sequence of the β B chain. Sequencing of the β-globin gene confirmed the variant gene as being an allele of the HBBB locus having the AAG → AGG and the AAA → AGA mutations at codons 66 and 144, respectively, both corresponding to the Lys → Arg substitution. The intrinsic oxygen affinity of the variant Hb (log P 50 = 0.79 at pH 7.0) was found to be intermediate between that of the sheep Hb B (log P 50 = 0.92) and that of Cypriot mouflon ( O. a. ophion ) Hb (log P 50 = 0.53), the latter having only the Lys → Arg change at β144, whereas nearly no differences were observed in the presence of the Cl − physiological effector. Result supports the indication that Arg at β144 enhances the role of the ligand in decreasing oxygen affinity, this effect being partially counteracted when Arg is at β66. Data also shows that the Lys → Arg change at β66 is responsible for 1.49 fold reduction in the intrinsic oxygen affinity. This hitherto undescribed variant increases to seven the number of alleles at the sheep HBBB locus . Following the nomenclature used for human Hb variants, the new allele was termed as the Hb Chios or [β B 66(E10) Lys → Arg, 144(HC1)Lys → Arg], whereas the proposed genetic nomenclature of the locus is HBBK.

The potentiality of xenon's sensitivity to its local magnetic environment is thoroughly investigated to probe internal structural differences between pig and horse metmyoglobin (MMb). These MMb's differ by 14 amino acids. One of these, Ile142 in horse MMb, is located in the proximal cavity, which is the xenon-binding site in horse MMb, and is replaced by Met142 in pig MMb. Specific and non-specific xenon-protein interactions are investigated here by 129Xe NMR chemical shifts and relaxation rate in aqueous solutions of pig MMb as a function of the xenon and protein concentrations. The results are complemented with 129Xe NMR data of the isostructural carbonmonoxy myoglobin (COMb), with computational calculations in order to highlight the structural differences between the cavities, and 1H NMR spectra to test the dependence of the 1H chemical shift on the addition of xenon. The 129Xe chemical shift NMR parameters are analysed quantitatively in terms of a two-site model. Xenon forms a 1:1 complex with the protein, characterized by an equilibrium binding constant K=[Xe]in/([Xe]out[MMb]), and exchanges rapidly between a cavity within the protein (X(ein)) and all other environments (Xe(out)). A comparison of equilibrium constant, K (74 M(-1)) in pig and K (146 M(-1)) in horse, reveals differences in affinity of xenon to the interior of pig MMb. Changes in xenon binding in both pig and horse MMb are also pointed out by other experimental results, e.g. the difference in the estimated delta(in), which is shifted downfield in pig MMb and upfield in horse MMb, with respect to 129Xe in buffer solution; the xenon-iron distance, 7.4 A, which is longer in the pig than was found in the horse, 5.3 A.

The functional properties of haemoglobin from the Mediterranean whale Balaenoptera physalus have been studied as functions of heterotropic effector concentration and temperature. Particular attention has been given to the effect of carbon dioxide and lactate since the animal is specialised for prolonged dives often in cold water. The molecular basis of the functional behaviour and in particular of the weak interaction with 2,3-diphosphoglycerate is discussed in the light of the primary structure and of computer modelling. On these bases, it is suggested that the A2 (Pro--Ala) substitution observed in the beta chains of whale haemoglobin may be responsible for the displacement of the A helix known to be a key structural feature in haemoglobins that display an altered interaction with 2,3-diphosphoglycerate as compared with human haemoglobin. The functional and structural results, discussed in the light of a previous study on the haemoglobin from the Arctic whale Balaenoptera acutorostrata, give further insights into the regulatory mechanisms of the interactive effects of temperature, carbon dioxide and lactate.

The Gymnothorax unicolor hemoglobin system is characterized by two components, called cathodic and anodic on the basis of their isoelectric point, which were separated by ion-exchange chromatography. The oxygen-binding properties of the purified components were studied in the absence and presence of chloride and/or GTP or ATP in the pH range 6.5-8.0. Stripped cathodic hemoglobin showed a small reverse Bohr effect, high oxygen affinity, and low co-operativity; the addition of chloride only caused a small decrease in oxygen affinity. In the presence of GTP or ATP, the oxygen affinity was dramatically reduced, the co-operativity increased, and the reverse Bohr effect abolished. Stripped anodic hemoglobin is characterized by both low oxygen affinity and co-operativity, and displayed a normal Bohr effect; the addition of chloride increased co-operativity, whereas ATP and GTP significantly modulated oxygen affinity at acidic pH values, enhancing the Bohr effect and giving rise to the Root effect. The complete amino-acid sequences of the alpha and beta chains of both hemoglobins were established; the molecular basis of the functional properties of the hemoglobins is discussed in the light of the primary structure and compared with those of other fish hemoglobins.

A study was made of the haemoglobin (Hb) system from the Sardinian dwarf horse (Equus caballus jara), one of the last surviving wild horse species in Europe. The oxygen binding properties of the whole haemolysate and of the four different horse Hbs, separated by ion-exchange chromatography, were studied with special regard to the effect of chloride, 2,3-diphosphoglycerate and lactate. Results indicate that no significant functional differences exist between the four Hb components of horse haemolysate. Moreover, the molecular basis of the intrinsically low oxygen affinity and of the weak interaction of horse Hb with 2,3-diphosphoglycerate is discussed in the light of the primary structure of the molecule and of the results of a computer modelling approach. On these bases, it is suggested that the A1 (Thr--Ser) and A2 (Pro--Gly) substitutions observed in the beta chains from horse Hb may be responsible for the displacement of the A helix that is known to be a key structural feature of those Hbs that display an altered interaction with 2,3-diphosphoglycerate as compared with human Hb.

The 13-globin gene cluster of the domestic sheep (Ovis aries) shows two common haplotypes: the A haplotype, which bears the adult sA allele (HBBA), and the B haplotype, which bears the adult sB allele (HBBB) [1, 2]. The chromosomal organization of the A haplotype was found to be similar to that present in goat (Capra hircus) since it shows the same triplication (5’ Er_Ea_NJsI sc_E[u_Erv_Jsrr_sn_£v_Evr_ NJs,rr-sF-3’) of an ancestral four gene set (E-E-NJs-s) [3]. This set is characterized by the presence of two embryonic genes (e), one pseudogene (NJs), and one adult gene (s). The expression of the adult gene varies during ontogenic development and under different physiological conditions. The sc, sA, and sF genes are, in fact, expressed during juvenile, adult and fetal life, respectively, and the sc gene expression is reactivable, at the expense of sA gene, under particular physiological or experimental conditions such as anemia and hypoxia or the administration of erythropoietin [4-6]. The B haplotype is considered to have diverged from the A haplotype, as the result of a recent deletion from a triplicated locus. In fact, due to the lack of the whole juvenile four-gene set containing the sc gene, it is duplicated (5’ er srr NJsr sB Errr-EIV yrsrI sF-3’) and sheep which are homozygous for the sB allele do not exhibit the sBia>scswitching

Haemocyanin from the lobster Palinurus elephas has been shown to change in its subunit composition according to the time of year. In contrast, in Palinurus mauritanicus, a lobster living at greater depth, no seasonal changes in subunit composition have been observed. The results obtained from a set of experiments performed on some Palinurus mauritanicus acclimatized in an aquarium have clearly indicated that modifications of haemocyanin subunit composition may be involved in the adaptation of arthropods to environmental change.

A study of the functional properties of haemoglobin from red deer (Cervus elaphus) whose habitat varies over a wide range of latitude, was performed. The oxygen-binding properties of the most common haemoglobin phenotype from the species living in Sardinia were examined with particular attention to the effect of pH, chloride, 2, 3-bisphosphoglycerate and temperature. Results indicate that red deer haemoglobin, like all haemoglobins from ruminants so far examined, is characterized by a low intrinsic oxygen affinity, with chloride being its main physiological modulator in vivo. The functional results and the low temperature sensitivity of the oxygen affinity are discussed in the light of the amino acid sequence of closely related ruminant haemoglobins.

We report the isolation and the functional characterization of alpha and beta chains from pig (Sus scropha domesticus) haemoglobin, as well as of the pig-human hybrid haemoglobins, alpha2(h)beta2(p) and alpha2(p)beta2(h) (i.e. Circe's haemoglobins), obtained by mixing the purified alpha and beta pig chains respectively with the corresponding partner human chains. Their functional properties have been compared with those of both parental haemoglobins in order to obtain information on the role of the different subunits and of their inter-relationships, both at the structural and functional levels. The results indicate that the functional properties of both hybrids are closer to those of the parental haemoglobin that provides the beta chains, confirming the major role of the beta chains in determining the oxygen affinity and the modulation mechanisms of the tetrameric molecule. This is supported by the thermodynamic properties, since the very low DeltaH of oxygen binding that characterizes pig haemoglobin and the alpha2(h)beta2(p) hybrid haemoglobin may be taken as the reflection of specific structural properties of pig beta chain.

Using capillary electrophoresis (CE) on a set of 21 peptides with a molecular mass ranging from about 350 to 1850 Da, the Stokes radii at different protonation stages and the acidic dissociation constants in water and in a 2,2,2-trifluoroethanol (TFE) water mixture (30% v/v) were determined. These results permitted us to establish separately the reliability of semiempirical models utilized for the prediction of peptide size and charge at different acidic pHapp (pHapp range: 2.00-4.25). The data obtained on size and charge were utilized in order to provide suitable mobility predictions on the basis of the charge-to-size ratio. The best predictive conditions for size and charge were found at the most acidic range of pHapp studied (2.00-2.25), either in water or a TFE-water mixture, and reliable predictive equations for peptide mobility were established at this pHapp.

The functional properties of Hb B of the wild European mouflon (Ovis gmelini musimon), Hb B of domestic sheep (Ovis aries), and Hb C isolated from anemic mouflon were investigated. Mouflon and sheep Hbs appear to be very similar in their response to organic anions and protons, whereas sheep Hb B displays an oxygen affinity lower than that of mouflon Hb B and sheep Hb A. Mouflon Hb B and Hb C, like sheep Hb A and Hb C, have similar efficiencies in transporting oxygen to the tissues. As in other ruminant Hbs, the effect of temperature on the oxygen affinity is slight. Data suggest that mouflon Hb B is not only structurally, but even functionally, more similar to sheep Hb A than to sheep Hb B.

Concerning the number and type of the hemoglobin components, the moray Muraena helena is characterized by three different phenotypes whose frequencies are nearly identical. Thus, the cathodal component is present in all individuals, whereas one or both of two anodal components may be present in the same phenotype. These components have been separated by chromatography. The oxygen binding properties of the purified hemoglobin components have been studied in the absence and presence of saturating concentrations of ATP or GTP and as a function of pH. The cathodal component shows an intrinsic O2 affinity four times higher than that of both anodal components, a very small Bohr effect and a significant decrease in O2 affinity upon addition of ATP and GTP (three and four times respectively with respect to stripped conditions), the latter being more effective than the former over the entire pH range examined. The anodal components do not appear functionally distinguishable and show the presence of an enhanced Bohr effect (Root effect) that is under the strict control of nucleotide triphosphates ATP, GTP, which, unlike in the cathodic component, exert the same effect on oxygen affinity. The complete sequence of the β chains of the cathodal and of one of the anodal components have been determined. The possible molecular basis of these different functional characteristics are discussed in the light of the globin sequence and of those amino acid residues which are known to be responsible of hemoglobin functional behaviour.

1. 1. The ligand-binding properties of hemoglobins from the urodelan amphibians Salamander salamander and Hydromantes genei genei have been studied and compared with those of hemoglobins from the urodelan, Triturus cristatus (Condo et al ., 1980; Condo et al ., 1983). 2. 2. Depending on the experimental conditions, the Bohr effect is absent or reversed in S. salamander hemoglobin, similarly to almost all the known hemoglobins from the Urodela (and tadpoles of both Anurans and the Urodela). 3. 3. H. genei shares, with anuran amphibia, a normal Bohr effect (reduced oxygen affinity of hemoglobin at acid pH), although smaller than that of mammals. 4. 4. The difference in the Bohr coefficient of hemoglobin from members of the two major subclasses of amphibians (Anura and Urodela: −0.3 and + 0.1, respectively, averaged from data of Table 1), almost vanishes in the whole blood (−0.2 to −0.4 in the two groups), due to the presence of intracellular effectors. 5. 5. Biochemical and physiological implications of the (apparent) reverse Bohr effect are reviewed.

1. 1. Hemocyanin from the mediterranean lobster Palinurus elephas has been shown to change in subunit composition with relation with the sex of the animal and the period of the year. 2. 2. The sexual difference, consisting of the presence of a specific subunit in the hemocyanin from female animals, is not linked to any evident functional property of the pigment. 3. 3. On the contrary, the season dependent variations in the subunit composition, which consist of the appearance of several catodic bands on electropherograms from both male and female animals, produce obvious functional effects; in particular an increase of the cooperativity of oxygen binding has been consistently found during the summer. 4. 4. The concentration of the pigment is also season and sex dependent, being higher in male animals and during the winter. 5. 5. Hemocyanin from a closely related lobster, Palinurus mauritanicus, does not show sexual and seasonal differences in subunit composition. This difference is tentatively attributed to the difference in the habitat of the two species.

The functional properties of the hemoglobin systems from two different species of bat i.e. Rousettus aegyptiacus and Myotis myotis have been studied as a function of chloride, polyphosphates, pH and temperature. Apart from overall similarities shared with most mammalian hemoglobins, the two systems show significant differences with respect to the effect of chloride and temperature sensitivity. These findings have been related to the different physiological needs of the two species.

The present study reports that long-term (18 days) administration of imipramine (IMI, 20 mg/kg) or desipramine (DMI, 15 mg/kg) produced a significant decrease in the GABA-stimulated 36Cl- uptake into membrane vesicles from the cerebral cortex of rats (experiment 1). Experiments 2A, B show that anti-immobility effects of DMI and IMI (subacute treatment) in the forced swimming test are enhanced when a single subconvulsant injection of picrotoxin or pentylenetetrazol is administered to the animals concurrently to the last antidepressant injection. These results are discussed in relation with a current GABAergic hypothesis of depression and antidepressant drug action.

Traitement des convulsions induites par le pentetrazol chez la souris par des antidepresseurs tricycliques (Imipramine et desipramine). Comparaison entre traitement chronique ou aigu, avant ou apres l'injection de pentetrazol