Localization of crustacean hyperglycemic hormone (CHH) in the X-Organ sinus gland complex in the eyestalk of the crayfish,Astacus leptodactylus (Nordmann, 1842)

The eyestalk of Astacus leptodactylus is investigated immunocytochemically by light, fluorescence, and electron microscopy, using an antiserum raised against purified crustacean hyperglycemic hormone (CHH). CHH can be visualized in a group of neurosecretory perikarya on the medualla terminalis (medulla terminalis ganglionic X-organ: MTGX), in fibers forming part of the MTGX-sinus gland tractus, and in a considerable part of the axon terminals composing the sinus gland. Immunocytochemical combined with ultrastructural investigations led to the identification of the CHH-producing cells and the CHH-containing neurosecretory granule type.     

The secretory dynamics of the CHH-producing cell group in the eyestalk of the crayfish,Astacus leptodactylus,in the course of the day/night cycle

Summary The secretory dynamics of the Crustacean Hyperglycemic Hormone (CHH)-producing cells in the eyestalk of the crayfish Astacus leptodactylus were studied during the daily cycle (12 h light/12 h dark). The different secretory stages of individual cells were determined by means of immunocytochemistry combined with morphometric analysis at the light-microscopic level. The data obtained were correlated with the 24-h rhythmicity of blood glucose concentration. The results suggest the following hypothesis. The synthetic activity of the CHH cells receives a stimulus 2 h before the beginning of the dark period, resulting in a pronounced transfer of CHH granules into the axons. These CHH granules reach the axon terminals after the onset of the dark period. At that time a burst of exocytotic activity occurs, causing a strong release of CHH into the hemolymph. Four hours later this CHH release results in hyperglycemia. The same process, though with less intensity, is repeated and causes a second smaller glucose peak at the beginning of the light period.     

DIATOM MINERALIZATION OF SILICIC ACID. II. REGULATION OF Si (OH)4 TRANSPORT RATES DURING THE CELL CYCLE OF NAVICULA PELLICULOSA1

Synchronized populations of Navicula pelliculosa (Bréb.) Hilse show a 10-fold increase in Si(OH)₄ transport rate during traverse through the cell division cycle. The transport activity pattern is similar to a “peak enzyme.” Kinetic analysis showed there was a significant change in K_s values, indicating increased “affinity” for Si(OH)₄ as cells neared maximal uptake rates. However, the dramatic changes in transport rate at various cell cycle stages were also reflected by alterations in the V_max, values of the transport process, suggesting a change in the number of functional transport “sites” in the plasma membrane. Cells in the wall forming stage, arrested from further development by Si(OH)₄ deprivation, maintained high transport rates for as long as 7 h. The rates decreased rapidly if protein synthesis were blocked or if Si(OH)₄ was added, the latter allowing the cells to traverse the rest of the cycle. The half-life of the transport activity ranged from 1.0 to 2.2 h when protein synthesis was inhibited at various cell cycle stages and during the natural decline of activity late in the cycle. The transport system appears to be metabolically unstable as is typical for a “peak protein.” The rise in transport rate through the cell cycle did not depend on the presence of Si(OH)₄ in the medium; therefore, the transport system does not appear to be induced by its substrate. The rise in transport is also observed in L:D synchronized cells developing in the presence of Si(OH)₄; neither does the transport system appear to be derepressed. The transport rate was strongly cell cycle-stage dependent; the data appeared to fit the “dependent pathway” model proposed by Hart-well to explain oscillations in enzyme synthesis during the cell cycle.     

Autoradiographic localization of opioid binding sites combined with immunogold detection of Leu-enkephalin,crustacean hyperglycaemic hormone and moult inhibiting hormone at the electron microscopic level in the sinus gland of the shore crab,Carcinus maenas

Double labelling experiments were performed on the same tissue section at the electron microscopic level, in order to show the involvement of the opioid leucine-enkephalin (Leu-enk) in the modulation of crustacean hyperglycaemic hormone (CHH) mobilization. Both neuropeptides were stored in distinct axon terminals of the sinus gland ofCarcinus maenas. A post-embedding immunogold cytochemical technique for Leuenk, CHH and the CHH neurohormone related moult inhibiting hormone (MIH) was combined with a scintillator intensified autoradiographic method to demonstrate binding of the opioid antagonist [³H] naloxone. Ultrathin sections were successively incubated with antisera against Leu-enk, CHH or MIH, and the corresponding colloidal gold labelled antisera, followed by autoradiographic processing. At the ultrastructural level [³H] naloxone binding sites were easily recognized by their silver tracks after development. Opioid binding sites for [³H] naloxone were visualized only at membranes of CHH-containing axon terminals. These results provide morphological evidence for direct enkephalinergic control of CHH containing neurons in the sinus gland ofC. maenas and are furthermore the first autoradiographic demonstration of opioid binding sites in the nervous system of invertebrates.     

Purification,characterisation and amino acid composition of the putative moult-inhibiting hormone (MIH) ofCarcinus maenas (Crustacea,Decapoda)

Summary Using a Y-organ in vitro assay to measure repression of ecdysteroid synthesis in the presence of putative moult-inhibiting hormone (MIH), in conjunction with HPLC separation of sinus gland neuropeptides ofCarcinus maenas, it was found that both the hyperglycemic hormone (CHH) and a novel peptide (argued to represent the MIH) inhibited ecdysteroid synthesis. The latter was purified to homogeneity, and amino acid analysis showed that it is a 61 residue peptide (minimum molecular mass 7,200 Da) with the following amino acid composition: Asx₉; Thr₂; Ser₂; Glx₇; Pro₁; Gly₄; Ala₂; 1/2 Cys₄; Val₄; Met₁; Ile₃; Leu₅; Tyr₁; Phe₃; His₃; Trp₂; Lys₂; Arg₆. The N-terminus appears to be blocked. MIH is at least 20 times more potent than CHH in repressing ecdysteroid synthesis and is active at concentrations of less than 250 pmol/l. There may be structural similarities between CHH and MIH, howeve, MIH displays no CHH radioimmunoreactivity or hyperglycemic activity. The physiological significance of CHH in controlling ecdysteroid titres is not known.Abbreviations CHH hyperglycemic hormone - MIH moult inhibiting hormone - PAGE polyacrylamide gel electrophoresis - RIA radioimmunoassay - SDS sodium dodecyl sulfate - SG smus gland(s) - SGE sinus gland equivalent - TFA trifluoroacetic acid     

Expression of recombinant eyestalk crustacean hyperglycemic hormone from the tropical land crab, <Emphasis Type="Italic">Gecarcinus lateralis</Emphasis>, that inhibits Y-organ ecdysteroidogenesis in vitro

Crustacean hyperglycemic hormone (CHH) is a pleiotropic neuropeptide that regulates carbohydrate and lipid metabolism, molting, reproduction, and osmoregulation in decapod crustaceans. CHH elevates glucose levels in the hemolymph by stimulating glycogenolysis in target tissues. It also inhibits ecdysteroidogenesis in the molting gland, or Y-organ (YO), possibly as a response to environmental stress. CHH acts via binding to a membrane receptor guanylyl cyclase, which is expressed in most tissues, including the YO. Large amounts of biologically active neuropeptide are required to investigate the mechanism of CHH signaling in the YO. Consequently, the eyestalk ganglia CHH (EG-CHH) isoform was cloned into a yeast (Pichia pastoris) expression vector to express recombinant mature peptide (rEG-CHH) with or without a C-terminal c-Myc/polyhistidine tag. Yeast cultures with untagged or tagged rEG-CHH inhibited ecdysteroidogenesis in YOs from European green crab (Carcinus maenas) 36% (P < 0.002) and 51% (P < 0.006), respectively. Purified tagged EG-CHH inhibited YO ecdysteroidogenesis 32% (P < 0.002), but lacked hyperglycemic activity in vivo. This is the first report of recombinant EG-CHH inhibiting YO ecdysteroidogenesis. The data suggest that the tagged recombinant peptide can be used to elucidate the CHH signaling pathway in the crustacean molting gland.     

CIRCADIAN MODULATION OF CRUSTACEAN HYPERGLYCEMIC HORMONE IN CRAYFISH EYESTALK AND RETINA

Previous studies suggested the retina could be a putative locus of daily crustacean hyperglycemic hormone (CHH) secretion, as it possesses its own metabolic machinery and is independent of the well-known CHH eyestalk locus responsible for the circadian secretion of this peptide. However, it has been proposed that hemolymph glucose and lactate concentrations play a dual role in the regulation of CHH in crayfish. To elucidate the temporal relationship between these two different CHH production loci and to examine their relationship with glucose regulation, we investigated the expression of CHH daily and circadian rhythms in the eyestalk and retina of crayfish using biochemical methods and time series analysis. We wanted to determine whether (1) putative retina and eyestalk CHH rhythmic expressions are correlated and if the oscillations of the two metabolic products of lactate and glucose in the blood due to CHH action on the target tissue correlate, and (2) retina CHH (RCHH) and the possible retinal substrate glycogen and its product glucose are temporally correlated. We found a negative correlation between daily and circadian changes of relative CHH abundance in the retina and eyestalk. This correlation and the cross-correlation values found between eyestalk CHH and hemolymph and glucose confirm that CHH produced by the X-organ sinus gland complex is under the previously proposed dual feedback control system over the 24?h time period. However, the presence of both glycogen and glucose in the retina, the cross-correlation values found between these parameters and hemolymph lactate and glucose, as well as RCHH and hemolymph and retina metabolic markers suggest RCHH is not under the same temporal metabolic control as eyestalk CHH. Nonetheless, their expression may be linked to common rhythms-generating processes. (Author correspondence: mlfm@hp.fciencias.unam.mx; mfajul@gmail.com)     

Variation in plasma glucose and pancreatic β cells in the turtle, Lissemys punctata (order: Chelonia; family: Trionychidae)

The present study relates to the determination of the plasma glucose level and volumetric analysis of β cells in pancreatic islets of the soft‐shelled turtle Lissemys punctata during different phases of its reproductive cycle. Reproductive events play a vital role in influencing the plasma glucose level and β‐cell behaviour in the pancreatic islets. The colour of the pancreas is either yellowish or pinkish, depending on endocrine activity. Islets are present throughout the gland and range from individual cells to small or large clumps, depending on the seasonal cycle. Splenic islets are dense with more blood capillaries and nerve innervations irrespective of sex and season. The endocrine cell mass forms irregular patches without connective tissue capsule. β cells occupy the inner region of the islets, being surrounded by other cell types. Lissemys punctata exhibits higher β‐cell activity during hibernation. Most insulin‐secreting cells acquire a larger size during the regressive period. An analysis indicates that β cells outnumber the non‐β endocrine cell mass in both number and per cent volume. There is negative correlation between islet mass and animal weight. Between the periods of reproductive cycles, a difference exists with respect to fasting plasma glucose and β‐cell volume.     

Expression of Biologically Active Crustacean Hyperglycemic Hormone (CHH) of <Emphasis Type="Italic">Penaeus monodon</Emphasis> in <Emphasis Type="Italic">Pichia pastoris</Emphasis>

Crustacean hyperglycemic hormone (CHH), molt-inhibiting hormone (MIH), and gonad-inhibiting hormone (GIH) are members of a major peptide family produced from the X-organ sinus gland complex in the eyestalk of crustaceans. This peptide family plays important roles in controlling several physiologic processes such as regulation of growth and reproduction. In this study the complementary DNA encoding a peptide related to the CHH/MIH/GIH family (so-called Pem-CMG) of the black tiger prawn Penaeus monodon was successfully expressed in the yeast Pichia pastoris under the control of the AOX1 promoter. The recombinant Pem-CMG was secreted into the culture medium using the -factor signal sequence; of Saccharomyces cerevisiae without the Glu-Ala-Glu-Ala spacer peptide. The amino terminus of the recombinant Pem-CMG was correctly processed as evidenced by amino-terminal peptide sequencing. The recombinant Pem-CMG was purified by reverse-phase high-performance liquid chromotography and used in a biological assay for CHH activity. The final yield of the recombinant Pem-CMG after purification was 260 µg/L of the culture medium. Both crude and purified recombinant Pem-CMG produced from P. pastoris showed the ability to elevate the glucose level in the hemolymph of eyestalk-ablated P. monodon, which demonstrates that Pem-CMG peptide functions as hyperglycemic hormone in P. monodon.     

Hyperglycemic responses to cold shock in the freshwater giant prawn, Macrobrachium rosenbergii

Crustacean hyperglycemic hormone (CHH), a neurohormone synthesized and released from the x-organ sinus gland complex, is primarily involved in carbohydrate metabolism; biogenic amines and peptidergic neuroregulators are known to modulate the release of CHH. Marked elevations of hemolymph glucose titers, which peaked within 2 h, were observed in both intact and bilaterally eyestalk-ablated prawns, Macrobrachium rosenbergii, when they were transferred directly from their optimal temperature of 28 °C to lower temperatures close to their lethal limit. Hyperglycemia can therefore be considered a characteristic response in this species under cold shock. Involvement of biogenic amines in the hyperglycemic response was also demonstrated. Hyperglycemic effects of epinephrine, dopamine and serotonin were mediated through CHH at the eyestalk level, but the response under cold shock was not exclusively mediated through CHH. It is suggested that factor(s) other than CHH are involved in the hyperglycemic response, possibly norepinephrine or/and octopamine. Accepted: 24 October 1998     

The specificity of epidermal chalone action: the results of in vivo experimentation with two purified skin extracts.

To date two inhibitors of epidermal cell proliferation have been characterized: (1) a factor which depresses DNA synthesis, and (2) a factor which depresses mitotic rate. In the absence of experimental proof it has been assumed that the respective targets for these purified inhibitory factors are in G₁ and G₂ phases of the cell cycle. In the experiments reported here both these fractions were subjected to cell cycle phase specificity tests in order to verify these assumptions. In addition, an epidermally derived “cell line” (the sebaceous gland) and two nonectodermal tissues were examined for a response. The results suggest that the response induced by the inhibitor of DNA synthesis is cell cycle phase-specific, that the target cells are at the G₁-S phase boundary, and that only epidermal cells respond. Similarly the factor which depresses the flow of cells from G₂ into mitosis had no measurable effect on DNA synthesis by any of the tissues tested. The G₂ inhibitor lacks an inhibitory effect on mitosis in the sebaceous gland.The physiological roles which epidermal chalones may play are briefly discussed. It is suggested that a G₁–G₂ chalone system may have been effective in isolating kinetically cell populations with modified function during the evolutionary development in the vertebrates.     

Quantitation of peptide hormone in single cultured secretory neurons of the crab,Cardisoma carnifex

The content of crustacean hyperglycemic hormone (CHH) in single cultured neurons of the crab Cardisoma carnifex was determined by a sensitive enzyme-linked immunosorbent assay (ELISA), using purified CHH (1–50 pg) of the crab Carcinus maenas as standard. The somata were dissociated from the group of 150 peptidergic neurons that form the X-organ—sinus gland neuroendocrine system. As previously reported, the neurons show immediate regenerative outgrowth in defined culture conditions, and develop, generally, into one of two morphological types: cells that produce broad, lamelliform growth cones (veils), and others that are characterized by branching of neurites. In this study, all but one of 64 veiling cells taken after various times in culture up to 12 days contained CHH. They could be readily categorized as having high (>33 pg; mean 86±5, S.E., n=47) or low (33pg; mean 22±2.5; n=17) Carcinus CHH equivalents. Thus, CHH is associated with neurons showing veiling outgrowth, but veiling neurons with low CHH form a distinct, but not morphologically distinguishable group. They may contain an isoform of CHH with limited cross-reactivity. In 24 branching neurons assayed, Carcinus CHH equivalents averaged 7.2±2 pg. This figure includes 14 neurons in which CHH was undetectable, and one that had 40 pg of Carcinus CHH equivalents. There was no significant change of the hormone content in cells of either type during 6 days of culturing.     

A comparative immunocytochemical investigation of the crustacean hyperglycemic hormone (CHH) in the eyestalks of some decapod crustacea

The eyestalk of the astacideans Orconects limosus, Nephrops norvegicus, and Homarus gammarus, and the palinuran Palinurus vulgaris, was examined with an antiserum raised against purified crustacean hyperglycemic hormone (CHH) of the astacidean species Astacus leptodactylus. A distinct immunopositive reaction occurs in a group of neurosecretory cells in the medulla terminalis ganglionic X-organ (MTGX), in the MTGX-sinus gland tractus, and in a considerable part of the sinus gland. The immunoreactive sites in the eyestalk of the investigated species correspond to the site of production, storage, and release of the CHH. Preliminary investigations with this antiserum also indicate that a positive immunoreaction can be obtained in the eyestalk of other decapod crustaceans, for example, of the brachyuran Macropipus puber and the caridean Palaemon serratus.     

Do the neurohormones VIH (vitellogenesis inhibiting hormone) and CHH (crustacean hyperglycemic hormone) of crustaceans have a common precursor? Immunolocalization of VIH and CHH in the X-organ sinus gland complex of the lobster,Homarus americanus

Summary

The present study deals with the location of the vitellogenesis inhibiting hormone (VIH)-producing cells in the eyestalk of the lobster Homarus americanus. In the present study, the neurosecretory pathways of VIH in Homarus, have been described immunocytochemically by use of a mouse serum against Homarus VIH. The location of the VIH cells was compared with the location of the crustacean hyperglycemic hormone (CHH) cells visualized by a rabbit serum raised against CHH of the crayfish Astacus leptodactylus. Immunocytochemical detection procedures, both at the light and electron microscopic level, revealed frequent but not complete co-localization of VIH and CHH in a variable number of the same group of perikarya. In the sinus gland, both neuropeptides were mostly demonstrated in distinct axonal endings characterized by different granule types. Postulations on the biosynthesis of these factors and suggestions concerning the processing of both neurohormones have been made.     

Regulation of circulating levels of the crustacean hyperglycemic hormone: evidence for a dual feedback control system

The effects of glutamate, aspartate, glycine, proline, alanine, taurine, glycerol, glucose and lactate injections on the haemolymph levels of the crustancean hyperglycemic hormone and/or glucose and lactate in the shore crab, Carcinus maenas, were investigated. Only glucose and lactate caused significant changes of hyperglycaemic hormone levels. Glucose injections resulted in a drop of both hormone and lactate, while lactate had an opposite effect, i.e. it raised both crustacean hormone and glucose levels. The results suggest that during increases in glycolytic flux, lactate may cause a release of hormone by a positive feedback mechanism. The hormone would then stimulate glycogenolysis, thus increasing glucose availability. If more glucose is released than is metabolized, excess glucose may leak from the cells and suppress crustancean hyperglycemic hormone release from the X-organ/sinus gland complex by negative feedback.Abbreviations ABTS 2,2-azino-bis (3-ethylbenzthiazoline sulphonic acid) - ANOVA one-way analysis of variance - BSA bovine serum albumin - BW body weight - CHH crustacean hyperglycemic hormone - ELISA cnzyme-liked immunosorbent assay - GIH gonadinhibiting hormone - IgG immunoglobin G - MIH moult-inhibiting hormone - MTGXO medulla terminalis X-organ - PB sodium phosphate buffer - PBS phosphate buffered saline - Pi inorganic phosphate - XO-SG X-organ-sinus gland complex     

Regulation of Enzyme Synthesis of the Glyoxylate,the Citric Acid,and the Methylcitric Acid Cycles in Candida lipolytica

Syntheses of the key enzymes of the glyoxylate cycle, in Candida lipolytica, were highly repressed by glucose. Syntheses of the key enzymes of the methylcitric acid cycle were also slightly repressed by glucose but the degrees of repression in the syntheses of these enzymes were nearly equal to those of repression in the syntheses of several enzymes of the citric acid cycle. All enzyme syntheses repressed by glucose were derepressed during incubation with succinate as well as with n-alkanes: enzyme syntheses of the methylcitric acid cycle did not necessitate the addition of propionate or odd-carbon n-alkanes. The enzymes of the methylcitric acid cycle seem to be constitutive, similarly as those of the citric acid cycle.

In the parent strain, the respective enzyme levels of the cells grown on an odd-numbered n-alkane were similar to those of the cells grown on an even-numbered n-alkane. But in the mutant strain lacking 2-methylisocitrate lyase, the cells grown on the odd-numbered alkane contained aconitate hydratase, NADP-Iinked isocitrate dehydrogenase, isocitrate lyase, 2- methylcitrate synthase and 2-methylaconitate hydratase all at higher levels than the cells grown on the even-numbered alkane. Both the parent cells and the mutant cells grown on the same carbon source contained at individually similar levels of the following six enzymes; citrate synthase, NAD-linked isocitrate dehydrogenase, succinate dehydrogenase, fumarate hydratase, malate dehydrogenase, and malate synthase. The pleiotropic changes of enzyme activities in the mutant cells grown on the odd-numbered alkane seem to be ascribable to direct or indirect stimulation caused by threo-d_s-2-methylisocitric acid accumulation.     

Assisted reproductive techniques with congenital hypogonadotropic hypogonadism patients: a systematic review and meta-analysis

Background

After hormonal replacement therapy (HRT) including androgen replacement or sequential therapy of estrogen and progesterone, The combination of human chorionic gonadotropin (hCG) and human menopausal gonadotropin (hMG) and pulsatile GnRH, is not sufficient to produce sufficient gametes in some patients with Congenital hypogonadotropic hypogonadism (CHH). A Systematic review and meta-analysis was performed to determine that assisted reproductive techniques (ART) can effectively treat different causes of infertility.

Methods

To determine the effect of ART on fertility of CHH patients and investigate whether outcomes are similar to infertility due to other causes, we conducted a systematic review and meta-analysis of retrospective trials.Clinical trials were systematically searched in Medline, Embase, and the Cochrane central register of controlled trials databases. The keywords and major terms covered “hypogonadotropic hypogonadism”, “kallmann syndrome”, “assisted reproductive techniques”, “intrauterine insemination”, “intracytoplasmic sperm injection”, “testicular sperm extraction”, “in vitro fertilization”, “embryo transplantation” and “intra-Fallopian transfer”.

Results

A total of 388 pregnancies occurred among 709 CHH patients who received ART (effectiveness 46, 95% confidence interval 0.39 to 0.53) in the 20 studies we included. The I² in trials assessing overall pregnancy rate (PR) per embryo transfer (ET) cycle was 73.06%. Similar results were observed in subgroup analysis by different gender. Regression indicates pregnancy rate decreases with increasing age. Fertilization, implantation and live birth rates (72, 36 and 40%) showed no significant differences as compared to infertility due to other causes.

Conclusions

Despite CHH patients usually being difficult to generate gametes, their actual chances of fertility are similar to subjects with other non-obstructive infertility. ART is a suitable option for CHH patients who do not conceive after long-term gonadotropin treatment.

     

GROWTH AND CELL CYCLE OF TWO CLOSELY RELATED RED TIDE-FORMING DINOFLAGELLATES: GYMNODINIUM NAGASAKIENSE AND G. CF. NAGASAKIENSE1

Small-sized vegetative cells were found to co-occur with normal-sized cells in populations of the European bloom-forming dinoflagellate Gymnodinium cf. nagasakiense Takayama et Adachi, currently known as Gyrodinium aureolum Hulburt, but not in populations of the closely related Japanese species Gymnodiniumn agasakiense. We examined how cell size differentiation may influence growth and cell cycle progression under a 12:12-h light: dark cycle in the European taxon, as compared to the Japanese one. Cell number and volume and chlorophyll red fluorescence in both species varied widely during the photocycle. These variations generally appeared to be related lo the division period, which occurred at night, as indicated by the variations of the fraction of binucleated cells (mitotic index) as well as the distribution of cellular DNA content. “Small” cells of G. cf. nagasakiense divided mainly during the first part of the dark period, although a second minor peak of dividing cells could occur shortly before light onset. In contrast, “large” cells displayed a sharp division peak that occurred 9 h after the beginning of the dark period. The lower degree of synchrony of “small” cells could be a consequence of their faster growth. Alternatively, these data may suggest that cell division is lightly controlled by an endogenous clock in “large” cells and much more loosely controlled in “small” cells. Cells of the Japanese species, which were morphologically similar to “large” cells of the European taxon, displayed an intermediate growth pattern between the two cell types of G. cf. nagasakiense, with a division period that extended to most of the dark period.     

Appearance and innervation of CHH-producing cells in the eyestalk of the crayfish Astacus leptodactylus examined after tracing with Lucifer Yellow

Summary By injection of the fluorescent dye Lucifer Yellow into individual Crustacean Hyperglycemic Hormone (CHH)-producing cells, the shape of these neurosecretory cells in the eyestalk of the crayfish Astacus leptodactylus can be traced. A highly fluorescent perikaryon gives rise to an axon that can be followed by the fluorescent label to the neurohemal region, the sinus gland. The proximal part of that axon sends out extensive branches into the neuropil of the medulla terminalis. Electron-microscopic investigations reveal synaptic input to these axonal ramifications.     

Glucose metabolism by trout (Salmo trutta) red blood cells

Summary Glucose metabolism has been studied in Salmo trutta red blood cells. From non-metabolizable analogue (3-O-methyl glucose and l-glucose) uptake experiments it is concluded that there is no counterpart to the membrane transport system for glucose found in mammalian red blood cells. Once within the cells, glucose is directed to CO₂ and lactate formation through both the Embden-Meyerhoff and hexose monophosphate shunts; lactate appears as the most important endproduct of glucose metabolism in these cells. From experiments under anaerobic conditions, and in the presence of an inhibitor of pyruvate transfer to mitochondria, most of the CO₂ formed appears to derive from the hexose monophosphate pathway. Appreciable O₂ consumption has been detected, but there is no clear relationship between this and substrate metabolism. Key enzymes of glucose metabolism hexokinase, fructose-6-phosphate kinase and, probably, pyruvate kinase are out of equilibrium, confirming their regulatory activity in Salmo trutta red blood cells. The presence of isoproterenol, a catecholamine analogue, induces important changes in glucose metabolism under both aerobic and anaerobic conditions, and increases the production of both CO₂ and lactate. From the data presented, glucose appears to be the major fuel for Salmo trutta red blood cells, showing a slightly different pattern of glucose metabolism from rainbow trout red blood cells.Abbreviations EM Embden-Meyerhoff pathway - G6D glucose-6-phosphate dehydrogenase - GOT glutamate oxalacetate transaminase - GPI glucose phosphate isomerase - HK hexokinase - HMS hexose monophosphate shunt - IP isoproterenol - LDH lactate dehydrogenase - MCB modified Cortland buffer - OMG 3-O-methyl glucose - PFK fructose-6-phosphate kinase - PK pyruvate kinase - RBC red blood cells - TAC tricarboxylic acid cycle