Defective remethylation of homocysteine is related to decreased synthesis of coenzymes B2 in thyroidectomized rats

Summary. We investigated the influence of hypothyroidism on homocysteine metabolism in rats, focusing on a hypothetical deficient synthesis of FAD by riboflavin kinases. Animals were allocated in control group (n=7), thyroidectomized rats (n=6), rats with diet deficient in vitamin B2, B9, B12, choline and methionine (n=7), thyroidectomized rats with deficient diet (n=9). Homocysteine was decreased in operated rats (2.6±1.01 vs. 4.05±1.0mol/L, P=0.02) and increased in deficient diet rats (29.56±4.52 vs. 4.05±1.0mol/L, P=0.001), when compared to control group. Erythrocyte-Glutathione-Reductase-Activation-Coefficient (index of FAD deficiency) was increased in thyroidectomized or deficient diet rats (P=0.004 for both). Methylenetetrahydrofolate-reductase and methionine-synthase activities were decreased in thyroidectomized rats but not in those subjected to deficient diet. Cystathionine--synthase was increased only in operated rats. Taken together, these results showed a defective re-methylation in surgical hypothyroidism, which was due in part to a defective synthesis of vitamin B2 coenzymes. This defective pathway was overcompensated by the increased Cystathionine--synthase activity.     

Augmented taurine release is not the mechanism of ischemic preconditioning’s cardioprotection

Summary. In ischemic preconditioning (IPC) a brief ischemic period protects the heart from a subsequent ischemic insult by an unknown mechanism. Osmotic swelling has been proposed to be a major cause of cell death when ischemic tissue is reperfused. The present study tests whether the preconditioned heart during reperfusion might release more taurine, an important osmolyte in the cardiac myocytes, to decrease cellular osmolarity, oppose swelling, and preserve viability. We collected the coronary effluent from isolated rabbit hearts for 10min before and 10min after preconditioning with 5min of global ischemia. The heart then experienced 15min of global ischemia and effluent was collected during reperfusion for 40min. A control group was studied similarly but without the preconditioning ischemia. Fifteen min of ischemia was chosen to avoid any taurine release caused by ischemic cell death. Taurine was measured with HPLC. In the IPC group there was a postischemic release over baseline of 5.09±1.51mol (approx 3.3% of the total taurine pool), whereas in the control group the release was not significantly different, 5.72±1.67mol. The percent of the taurine pool lost from each heart during reperfusion was calculated based on an assumption of a total content of 20M taurine/gm wet weight. Since the amount of taurine released by the isolated rabbit heart following ischemia was not different in preconditioned and non-preconditioned hearts, we conclude that reduced swelling through taurine release is not the mechanism of the cardioprotective effects of IPC.     

Evidence for expression of a single distinct form of mammalian cysteine dioxygenase

Summary. Cysteine dioxygenase (CDO) plays a critical role in the regulation of cellular cysteine concentration. Because multiple forms of CDO (23kDa, 25kDa, and 68kDa) have been claimed based upon separation and detection using SDS-PAGE/western blotting (with antibodies demonstrated to immunoprecipitate CDO), we further investigated the possibility of more than one CDO isoform. Using either rabbit antibody raised against purified rat liver CDO or against purified recombinant his₆-tagged CDO (r-his₆-CDO) and using 15% (wt/vol) polyacrylamide for the SDS-PAGE, we consistently detected the 25kDa band, but never detected a 68kDa band, in rat liver, kidney, lung and brain. Nondenatured gel electrophoresis of r-his₆-CDO yielded a molecular mass estimate of 25.7kDa and no evidence of dimerization. Mass spectrometry of r-his₆-CDO yielded two peaks with molecular masses of 24.1kDa and 24.3kDa. Anion-exchange FPLC of r-his₆-CDO also gave two peaks, with the first containing CDO that was 7.5-times as active as the more anionic form that eluted second. When the two peaks recovered from FPLC were run on SDS/PAGE, the first (more active) CDO fraction yielded two bands (perhaps as an artifact of SDS/PAGE), whereas the second (less active) CDO fraction yielded only the 23kDa band. We conclude that the physiologically active form of CDO is the 25kDa (i.e., 23.5kDa based on mass spectrometry) monomer and that this active form is probably derived by post-translational modification of the 23kDa gene product.     

Taurine protected myocardial mitochondria injury induced by hyperhomocysteinemia in rats

Summary. Taurine can protect against cardiovascular diseases, whereas elevated levels of plasma homocysteine are associated with atherosclerotic and thromboembolic cardiovascular diseases. To illustrate the effects of taurine on hyperhomocysteinemia, we observed the myocardial mitochondria dysfunction in the rats with hyperhomocysteinemia induced by diet methionine loading, and the therapeutic effect of taurine. A methionine diet increased plasma homocysteine concentration (133.51±27.91mol/L vs 12.31±2.58mol/L in control, P<0.01), stimulated the production of reactive oxygen species (ROS) in the myocardial mitochondria, and inhibited the activities of mitochondrial Mn-superoxide dismutase and catalase. The ⁴⁵Ca uptake and Ca²⁺-ATPase activity in the myocardial mitochondria were significantly lowered in rats with hyperhomocysteinemia. Taurine supplements effectively attenuated the hyperhomocysteinemia-induced ROS production and inhibition of Mn-superoxide dismutase and catalase activities in the myocardial mitochondria, and increased its ⁴⁵Ca uptake and Ca²⁺-ATPase activity. Thus, taurine antagonizes the oxidative stress injury in the myocardial mitochondria induced by the hyperhomocysteinemia.     

Betaine or taurine administration prevents fibrosis and lipid peroxidation induced by rat liver by ethanol plus carbon tetrachloride intoxication

Summary. The aim of this study was to investigate the effect of betaine or taurine on liver fibrogenesis and lipid peroxidation in rats. Fibrosis was induced by treatment of rats with drinking water containing 5% ethanol and CCl₄ (2×weekly, 0.2ml/kg, i.p.) for 4 weeks. Ethanol plus CCl₄ treatment caused increased lipid peroxidation and disturbed antioxidant system in the liver. Histopathological findings suggested that the development of liver fibrosis was prevented in rats treated with betaine or taurine (1% v/v in drinking water) together with ethanol plus CCl₄ for 4 weeks. When hepatic taurine content was depleted with -alanine (3% v/v in drinking water), portal-central fibrosis induced by ethanol+CCl₄ treatment was observed to proceed cirrhotic structure. Betaine or taurine was also found to decrease serum transaminase activities and hepatic lipid peroxidation without any change in hepatic antioxidant system in rats with hepatic fibrosis. In conclusion, the administration of betaine or taurine prevented the development of liver fibrosis probably associated with decreased oxidative stress.     

Long-term taurine supplementation reduces mortality rate in streptozotocin-induced diabetic rats

Summary. Oxidative stress is implicated in the pathogenesis of diabetes mellitus. Taurine and vitamin E+selenium supplementation has some benefits in experimental models of diabetes mellitus. This study evaluates whether taurine and vitamin E+selenium supplementations reduce a hard end-point such as mortality due to diabetes. Streptozotocin-induced diabetic rats were fed with standard diet or taurine (5%, w/w) or vitamin E (500UI/Kg)+selenium (8mg/Kg) enriched diets. Taurine significantly decreased mortality rate (p<0.04), while vitamin E failed to increase survival. In the late phase of the disease, taurine significantly decreased glycaemia, being vitamin E ineffective. No correlation between glycaemia and survival was found. None of supplementations modified body weight. Thus, only taurine decreases the mortality rate and glycaemia. These results encourage new research in the field, since classical hypoglycaemic agents are unable to decrease mortality in diabetic patients.     

Interaction of aluminium ions with some amino acids present in human blood

Summary. The interaction of aluminium with some amino acids present in human blood was studied combining ion-chromatography (IC), atomic absorption spectrometry (AAS) and ultrafiltration (UF) techniques. An IC system for simultaneous determination of ornithine, lysine, glutamic acid, aspartic acid and tyrosine was developed. By adding aluminium to standard solutions of the amino acids and keeping the pH at 6 and 7 it was possible to verify that aluminium caused a reduction on the amino acid chromatographic signals. Similar experiment, carried out for copper showed the same behaviour (with different percentage of signal reductions) and validated the results for aluminium, considering that the interaction Cu-amino acid is well-established. The AAS analysis of sample fractions (500l) after the IC separation showed that aluminium (as copper as well) is not present in the fractions in which the amino acid peaks appear in the chromatogram. These approaches carried out with serum samples after UF showed that part of the free fraction of serum aluminium is distributed, besides other ligands, among these amino acids. It was found that in serum the affinity for aluminium followed the sequence Lys>Orn>Tyr>GluAsp.     

Nefopam inhibits calcium influx, cGMP formation, and NMDA receptor-dependent neurotoxicity following activation of voltage sensitive calcium channels

Summary. Nefopam hydrochloride is a potent non sedative benzoxazocine analgesic that possesses a profile distinct from that of anti-inflammatory drugs. Previous evidence suggested a central action of nefopam but the detailed mechanism remains unclear. We have investigated the actions of nefopam on voltage sensitive calcium channels and calcium-mediated pathways. We found that nefopam prevented N-methyl-D-aspartate (NMDA)-mediated excitotoxicity following stimulation of L-type voltage sensitive calcium channels by the specific agonist BayK8644. Nefopam protection was concentration-dependent. 47M nefopam provided 50% protection while full neuroprotection was achieved at 100M nefopam. Neuroprotection was associated with a 73% reduction in the BayK8644-induced increase in intracellular calcium concentration. Nefopam also inhibited intracellular cGMP formation following BayK8644 in a concentration-dependent manner, 100M nefopam providing full inhibition of cGMP synthesis and 58M allowing 50% cGMP formation. Nefopam reduced NMDA receptor-mediated cGMP formation resulting from the release of glutamate following activation of channels by BayK8644. Finally, we also showed that nefopam effectively reduced cGMP formation following stimulation of cultures with domoic acid, while not providing neuroprotection against domoic acid. Thus, the novel action of nefopam we report here may be important both for its central analgesic effects and for its potential therapeutic use in neurological and neuropsychiatric disorders involving an excessive glutamate release.     

Characteristics of basal taurine release in the rat striatum measured by microdialysis

Summary. Taurine is a sulfur-containing amino acid thought to be an osmoregulator, neurotransmitter or neuromodulator in the brain. Our objective was to establish how much taurine is released in the striatum and examine the mechanisms controlling extracellular taurine concentrations under resting conditions. The experiments were made on rats by microdialysis in vivo. Changes in taurine were compared with those in glutamate, glycine and the non-neuroactive amino acid threonine. Using the zero net flux approach we showed the extracellular concentration of taurine to be 25.2±5.1M. Glutamate was increased by tetrodotoxin and decreased by Ca²⁺ omission, glycine and threonine were not affected and both treatments increased extracellular taurine. The basal taurine release was increased by the taurine transport inhibitor guanidinoethanesulfonate and reduced by the anion channel blocker 4-acetamido-4-isothiocyanatostilbene-2,2-disulfonic acid.     

N-Terminal pyrazinones: a new class of peptide-bound advanced glycation end-products

Summary. The reaction of peptide Gly-Ala-Phe with the -dicarbonyl compounds glyoxal and methylglyoxal was studied under physiological conditions (pH=7.4, 37°C). Using HPLC with UV and fluorescence detection, a rapid derivatization of the peptide and the concomitant formation of well-defined products were observed. The products, which showed characteristic UV absorbance (_max=320 to 340nm) and fluorescence (_ex=330 to 340nm, _em=395 to 405nm), were identified by ESI-MS and NMR spectroscopic analysis as the N-terminally pyrazinone-modified peptides I (N-[2-(2-oxo-2H-pyrazin-1-yl)-propyl]-phenylalanine) and II (N-[2-(5-methyl-2-oxo-2H-pyrazin-1-yl)-propionyl]-phenylalanine). Model experiments revealed that the reactivity of the N-termini of peptides towards a derivatization by glyoxal is in the same order of magnitude as that of arginine, which generally is attributed as main target for -dicarbonyl compounds in proteins. Incubation of insulin with glyoxal proved the protein-bound formation of pyrazinones, with the N-terminus of the B-chain as the main target. According to these results, we conclude that N-terminal pyrazinones represent a new type of advanced glycation end-products (AGEs) with significance for biological systems and foods.     

Cysteine-induced hypoglycemic brain damage: an alternative mechanism to excitotoxicity

Summary. Central neural damage caused by L-cysteine (L-Cys) was first reported more than 30 years ago. Nevertheless, the exact mechanisms of L-Cys-mediated neurotoxicity are still unclear. Preliminary study in mice demonstrated that, following L-Cys injection, animals developed tachypnea, tremor, convulsions, and death in conjunction with documented hypoglycemia. The aim of the present study was to further investigate the mechanism of L-Cys-mediated hypoglycemic effect and neural damage. Neonatal ICR mice (n=6) were injected with L-Cys (0.5–1.5mg/g body weight [BW]), and their blood glucose and insulin levels were determined up to 90min following the injection. Experiments were repeated in chemically (streptozotocin [STZ]) pancreatectomized animals. Brain histology was assessed. Mice injected with L-Cys exhibited dose-dependent neurotoxicity and higher mortality as compared with controls. L-Cys (1.2–1.5mg/g BW) caused severe hypoglycemia (glucose<42mg/dl) (P<0.001). In STZ-treated (diabetic) animals, L-Cys (1.5mg/g BW) increased plasma insulin levels 2.3-fold and decreased serum glucose levels by 50% (P<0.01). Brain histology revealed destruction of as much as 51% of hippocampal neurons in the L-Cys-treated mice but not in the glucose-resuscitated animals. These findings suggest that L-Cys injection can cause pronounced hypoglycemia and central neural damage which is glucose reversible. Since L-Cys is chemically different from the other excitatory amino acids (glutamate and aspartate), L-Cys-mediated neurotoxicity may be connected to its hypoglycemic effect.     

Role of taurine supplementation to prevent exercise-induced oxidative stress in healthy young men

Summary. To evaluate the protective effects of taurine supplementation on exercise-induced oxidative stress and exercise performance, eleven men aged 18–20 years were selected to participate in two identical bicycle ergometer exercises until exhaustion. Single cell gel assay (SCG assay) was used to study DNA damage in white blood cells (WBC). Pre-supplementation of taurine, a significant negative correlation was found between plasma taurine concentration before exercise and plasma thiobaribituric-acid reactive substance (TBARS) 6hr after exercise (r=–0.642, p<0.05). WBC showed a significant increase in DNA strand breakage 6hr and 24hr after exercise. Seven-day taurine supplementation reduced serum TBARS before exercise (p<0.05) and resulted in a significantly reduced DNA migration 24hr after exercise (p<0.01). Significant increases were also found in VO₂max, exercise time to exhaustion and maximal workload in test with taurine supplementation (p<0.05). After supplementation, the change in taurine concentration showed positive correlations with the changes in exercise time to exhaustion and maximal workload. The results suggest that taurine may attenuate exercise-induced DNA damage and enhance the capacity of exercise due to its cellular protective properties.     

Comparative residual toxicities of pesticides to the predator Agistemus industani (Acari: Stigmaeidae) on citrus in Florida

Residual toxicities of registered and selected experimental pesticides used on citrus against Agistemus industani Gonzalez (Acari: Stigmaeidae) were compared. Pesticides considered highly toxic to A. industani were: abamectin 0.15 EC at 731ml/ha+FC 435-66 petroleum oil at 46.8l/ha, pyridaben 75WP at 469g/ha, ethion 4EC at 7.01l/ha+FC 435-66 petroleum oil at 46.8l/ha, propargite 6.55 EC at 3.51l/ha, chlorfenapyr 2SC at 1.46l/ha applied alone or in combination with FC 435-66 petroleum oil at 46.8l/ha, sulphur 80DF at 16.81kg/ha, dicofol 4EC at 7.01l/ha, fenbutatin oxide 50WP at 2.24kg/ha, benomyl 50WP at 2.24kg/ha, benomyl 50WP at 1.68kg/ha+ferbam 76 GF at 5.60kg/ha, ferbam 76GF at 11.21kg/ha, neem oil 90EC at 46.8l/ha, and copper hydroxide DF (40% metallic copper) at 4.48kg metallic copper/ha+FC 435-66 petroleum oil at 46.8l/ha. Pesticides that were moderately to slightly toxic included: copper sulphate 98% at 4.48kg metallic copper/ha+FC 435-66 petroleum oil at 46.8l/ha, fenbuconazole 2F at 280ml/ha+FC 435-66 petroleum oil at 46.8l/ha, FC 435-66 petroleum oil applied alone at 46.8l/ha or 23.4l/ha, and diflubenzuron 25WP at 1.40kg/ha. Pesticides that were non-toxic included: fenbuconazole 2F at 585ml/ha, malathion 57EC at 5.85l/ha, FC 435-66 petroleum oil at 46.8l/ha, carbaryl 80S at 3.36kg/ha, chlorpyrifos 4EC at 4.68l/ha, and formetanate 92SP at 1.12kg/ha. Understanding the toxic effects of field weathered pesticides against key predacious mite species is important for effective IPM. The results of this study provide a comparison of direct and indirect toxic effects of various pesticides to A. industani under field conditions.     

Recombinant Components of the EcoRII Restriction–Modification System: Restriction Endonuclease Can Interact with DNA · RNA Duplexes

To obtain recombinant restriction endonuclease (R) and methylase (M) of the EcoRII restriction–modification system, bacterial strains overproducing their functional hexahistidine derivatives were constructed. Active full-length R·EcoRII was produced only in cells that also expressed M·EcoRII from a multicopy plasmid. Recombinant R·EcoRII bound with hybrid DNA·RNA duplexes.     

Proliferation and germination of somatic embryos from embryogenic suspension cultures in <Emphasis Type="Italic">Phœnix dactylifera</Emphasis>

The present study demonstrates a procedure for the rapid development of a high number of somatic embryos from embryogenic suspension culture. This method might be efficient for mass propagation of Phnix dactylifera L. Embryogenic callus placed in liquid medium with 10^–5M ABA yielded an average 72 embryos per 100ml of culture medium within 2months, while those placed on solid medium yielded an average of 33, 20 and 16 embryos per 100ml of culture medium respectively for 10^–7, 10^–6 and 10^–5 M ABA after 4months. The combination of 2,4-DIchlorophenoxyacetic acid (2,4-D) (4.5×10^–7M), glutamine (6.7×10^–4M), and ABA (10^–5M) (L8 liquid medium) showed a beneficial effect on somatic embryos production compared to 2,4-D and glutamine alone, while this combination significantly (p<0.05) increased the accumulation of storage proteins (144 and 138mgg^–1 DW respectively for Jihel and Bousthami noir cultivars) in somatic embryos. The somatic embryos which underwent maturation on medium containing only 4.5×10^–7M 2,4-D and 10^–5M ABA (L6 liquid medium) accumulated more sugars (292 and 265mgg^–1 DW respectively for Jihel and Bousthami noir) than those matured on any other liquid medium. Histological studies revealed that somatic embryos (developed in L6 and L8 liquid media) accumulated less reserve compounds (proteins and sugars) than zygotic embryos. The addition of activated charcoal (0.25 and 0.5gl^–1) and phytagel^® (2.5gl^–1) to the germination medium may be useful for enhancing the germination of Phnix dactyliferasomatic embryos.     

Macronutrient input from pollen in two regenerating pine stands in southeast Korea

This study examined macronutrient input from pollen in two naturally regenerating pine stands in southeast Korea. Durham gravity pollen collectors were used to measure pine pollen deposition and the macronutrients in the collected pine pollen were analyzed. In 1998, pine pollen deposition began just before 18 April and lasted for approximately 2weeks. Total pine pollen deposition differed between the two sampling sites; 27.5kgha^–1 was collected from the mature stand and 17.7kgha^–1 was collected from the young stand. The values for nutrient deposition from pine pollen are 549gha^–1 N, 78gha^–1 P, 240gha^–1 K, 45gha^–1 S and 22gha^–1 Mg at the mature stand and 353gha^–1 N, 51gha^–1 P, 151gha^–1 K, 27gha^–1 S and 14gha^–1 Mg at the young stand, suggesting that nutrients from pine pollen contribute to forest nutrient cycling. The pine pollen deposition values obtained from our study (17.7–27.5kg^–1ha^–1year^–1) are approximately 1/115–180-fold that of pine litterfall in Korea. If we take pollen nutrients into account, the contribution rate of pollen to the annual nutrient input is very high in our study (N 1/30, P 1/5, K 1/9 that of litterfall). Macronutrient deposition from pine pollen is concentrated temporally in spring. Although the annual contribution of nutrient mass by pollen is small compared to that of litterfall, the rapid turnover rate of pollen nutrients combined with episodic deposition suggests that pollen may play a disproportionate role in temperate pine forest nutrient cycling.     

Early development of the self-fertilizing mangrove killifish <Emphasis Type="Italic">Rivulus marmoratus</Emphasis> reared in the laboratory

The mangrove killifish Rivulus marmoratus was reared at 25°±1°C and 17ppt salinity from 0 to 100 days after hatching (DAH), and its early development was described by examining growth and morphometric parameters, meristic characters (vertebral and fin-ray counts), bone-cartilage development, and pigmentation. Growth was isometric for preanal length, head length, snout length, body depth, pectoral-fin length, dorsal-fin length, anal-fin length, and caudal-peduncle depth. Negative allometric growth was observed in eye diameter and gape size. Meristic counts (mean±SD) for vertebrae (34.2±0.4) and dorsal- (8.6±0.5), anal- (11.4±0.5), and caudal-fin rays (30.2±0.8) were complete at 0 DAH (n=5), whereas pectoral-fin rays and pelvic-fin rays were complete by 30 DAH (14.5±0.4, n=5) and 60 DAH (4.2±0.8, n=5). Full ossification of meristic elements proceeded in the following sequence: vertebrae (by 30 DAH), caudal-, dorsal-, and anal-fin rays (by 60 DAH), pectoral-fin rays (between 60 DAH and 100 DAH), and pelvic-fin rays (by 100 DAH). Both morphological characters and meristic counts indicate that this species can be considered to be a juvenile after 9.8mm in standard length (20 DAH).     

The North Sea: source or sink for nitrogen and phosphorus to the Atlantic Ocean?

Annual nitrogen and phosphorus budgets for the whole North Sea taking into account the most recent data available were established. The area considered has a total surface of approximately 700,000km² and corresponds to the definition by OSPARCOM (Oslo and Paris Commission) with the exclusion of the Skagerrak and Kattegat areas. Input and output fluxes were determined at the marine, atmospheric, sediment and continental boundaries, and riverine inputs based on river flows and nutrient concentrations at the river–estuary interface were corrected for possible estuarine retention. The results showed that the North Sea is an extremely complex system subjected to large inter-annual variability of marine water circulation and freshwater land run-off. Consequently, resulting total N (TN) and P (TP) fluxes are extremely variable from 1 year to another and this has an important influence on the budget of these elements. Total inputs to the North Sea are 8870±4860kTNyear^–1 and 494±279kTPyear^–1. Denitrification is responsible for the loss of 23±7% of the TN inputs while sediment burial is responsible for the retention of only of 2±2% of the TP input. For TN, due to the large variability on marine and estuarine fluxes, and to the uncertainty related to the denitrification rate, it was concluded that the North Sea could either be a source (1930kTNyear^–1) or a sink (1700kTNyear^–1) for the waters of the North Atlantic Ocean. For TP it was concluded that the North Sea is mostly a source (–4 to 52kTPyear^–1) for the waters of the North Atlantic Ocean.     

Diel Movements of Bat Rays, Myliobatis californica, in Tomales Bay, California: Evidence for Behavioral Thermoregulation?

We used ultrasonic telemetry to examine movement patterns of 11 bat rays, Myliobatis californica, in Tomales Bay, California. Tomales Bay is long (20km) and narrow (1.4km), and is hydrographically separated into outer and inner bay regions. The outer bay (the outermost 8km) is characterized by oceanic conditions while the shallow inner bay (the innermost 12km) features wide seasonal temperature shifts. Five rays were tracked monthly from October 1990 to November 1991 and six rays (four of which carried temperature-sensing transmitters) were tracked daily from 30 June to 16 July 1992. Mean bat ray movement rate was 8.84mmin^–1 (range 4.49 to 13.40mmin^–1) and was not significantly affected by size (p=0.592), tidal stage (p=0.610), or time of day (p=0.327). Movement direction was unrelated to tidal stage (p=0.472) but showed a highly significant diel pattern (p<0.001). From 2:50–14:50h, rays moved toward the warmer and shallower inner bay, while from 14:50–2:50h they moved toward the cooler and deeper outer bay. These telemetry data, along with known bat ray foraging patterns and respiratory temperature-sensitivity, argue for behavioral thermoregulation as the primary influence on this movement pattern.     

Flexibility on the karyotype evolution in bitterlings (Pisces, Cyprinidae)

In bitterlings (Acheilognathinae) C- and Ag-banding karyotypes of 6 species-subspecies collected in China and South Korea were analyzed. The chromosomal constitution of 2n=46 (4SM+42ST) in Rhodeus atremius fangi was quite different from that of 2n=48 (8M+20SM+20ST) in other species-subspecies in Rhodeus. It was concluded from the analysis of banded chromosomes that the increase in number of ST during the karyotype change from 2n=48 to 2n=46 was achieved by a series of pericentric inversions from 24 M-SM to 24 ST, and the decrease in the diploid number was caused by an additional tandem fusion of 4 ST chromosomes, forming a new ST pair in the 2n=46 karyotype. The karyotype of Tanakia koreensis, T. signifer, and Acheilognathus macropterus is 2n=48 (8M+20SM+20ST), 2n=48 (8M+20SM+14–16ST+4–6 A), 2n=44 (14M+16SM+14ST), respectively. In R. ocellatus ocellatus, T. koreensis, T. signifer and A. macropterus, karyotype changes from 2n=48 to 2n=44 due to centric fusion and inversion have also been estimated. It was suggested that C-banding heterochromatin was greatly concerned with the karyotype evolution in bitterlings.