Genetic divergence with ongoing gene flow is maintained by the use of different hosts in phytophagous ladybird beetles genus Henosepilachna.

Adaptation to different environments can promote population divergence via natural selection even in the presence of gene flow - a phenomenon that typically occurs during ecological speciation. To elucidate how natural selection promotes and maintains population divergence during speciation, we investigated the population genetic structure, degree of gene flow and heterogeneous genomic divergence in three closely related Japanese phytophagous ladybird beetles: Henosepilachna pustulosa, H. niponica and H. yasutomii. These species act as a generalist, a wild thistle (Cirsium spp.) specialist and a blue cohosh (Caulophyllum robustum) specialist, respectively, and their ranges differ accordingly. The two specialist species widely co-occur but are reproductively isolated solely due to their high specialization to a particular host plant. Genomewide amplified fragment-length polymorphism (AFLP) markers and mitochondrial cytochrome c oxidase subunit I (COI) gene sequences demonstrated obvious genomewide divergence associated with both geographic distance and ecological divergence. However, a hybridization assessment for both AFLP loci and the mitochondrial sequences revealed a certain degree of unidirectional gene flow between the two sympatric specialist species. Principal coordinates analysis (PCoA) based on all of the variable AFLP loci demonstrated that there are genetic similarities between populations from adjacent localities irrespective of the species (i.e. host range). However, a further comparative genome scan identified a few fractions of loci representing approximately 1% of all loci as different host-associated outliers. These results suggest that these three species had a complex origin, which could be obscured by current gene flow, and that ecological divergence can be maintained with only a small fraction of the genome is related to different host use even when there is a certain degree of gene flow between sympatric species pairs.

Beyond counts and shapes: studying pathology of dendritic spines in the context of the surrounding neuropil through serial section electron microscopy.

Because dendritic spines are the sites of excitatory synapses, pathological changes in spine morphology should be considered as part of pathological changes in neuronal circuitry in the forms of synaptic connections and connectivity strength. In the past, spine pathology has usually been measured by changes in their number or shape. A more complete understanding of spine pathology requires visualization at the nanometer level to analyze how the changes in number and size affect their presynaptic partners and associated astrocytic processes, as well as organelles and other intracellular structures. Currently, serial section electron microscopy (ssEM) offers the best approach to address this issue because of its ability to image the volume of brain tissue at the nanometer resolution. Renewed interest in ssEM has led to recent technological advances in imaging techniques and improvements in computational tools indispensable for three-dimensional analyses of brain tissue volumes. Here we consider the small but growing literature that has used ssEM analysis to unravel ultrastructural changes in neuropil including dendritic spines. These findings have implications in altered synaptic connectivity and cell biological processes involved in neuropathology, and serve as anatomical substrates for understanding changes in network activity that may underlie clinical symptoms.

Glutamate and GABA receptors and transporters in the basal ganglia: what does their subsynaptic localization reveal about their function?

GABA and glutamate, the main transmitters in the basal ganglia, exert their effects through ionotropic and metabotropic receptors. The dynamic activation of these receptors in response to released neurotransmitter depends, among other factors, on their precise localization in relation to corresponding synapses. The use of high resolution quantitative electron microscope immunocytochemical techniques has provided in-depth description of the subcellular and subsynaptic localization of these receptors in the CNS. In this article, we review recent findings on the ultrastructural localization of GABA and glutamate receptors and transporters in monkey and rat basal ganglia, at synaptic, extrasynaptic and presynaptic sites. The anatomical evidence supports numerous potential locations for receptor-neurotransmitter interactions, and raises important questions regarding mechanisms of activation and function of synaptic versus extrasynaptic receptors in the basal ganglia.

Genetic diagnosis of sex chromosome aberrations in horses based on parentage test by microsatellite DNA and analysis of X- and Y-linked markers.

REASONS FOR PERFORMING STUDY: Sex chromosome aberrations are often associated with clinical signs that affect equine health and reproduction. However, abnormal manifestation with sex chromosome aberration usually appears at maturity and potential disorders may be suspected infrequently. A reliable survey at an early stage is therefore required. OBJECTIVES: To detect and characterise sex chromosome aberrations in newborn foals by the parentage test and analysis using X- and Y-linked markers. METHODS: We conducted a genetic diagnosis combined with a parentage test by microsatellite DNA and analysis of X- and Y-linked genetic markers in newborn light-breed foals (n = 17, 471). The minimum incidence of sex chromosome aberration in horses was estimated in the context of available population data. RESULTS: Eighteen cases with aberrations involving 63,XO, 65,XXY and 65,XXX were found. The XO, XXY (pure 65,XXY and/or mosaics/chimaeras) and XXX were found in 0.15, 0.02 and 0.01% of the population, respectively, based solely on detection of abnormal segregation of a single X chromosome marker, LEX003. CONCLUSIONS AND POTENTIAL RELEVANCE: Detection at an early age and understanding of the prevalence of sex chromosome aberrations should assist in the diagnosis and managment of horses kept for breeding. Further, the parental origin of the X chromosome of each disorder could be proved by the results of genetic analysis, thereby contributing to cytogenetic characterisation.

[Adrenal gland lymphoma accompanied by renal cell carcinoma].

A 76-year-old man was admitted with right hypochondralgia. Computed tomography revealed tumors measuring 90 mm and 20 mm in the right adrenal gland and kidney, respectively. Under a diagnosis of renal and adrenal cancer, the patient underwent nephro-adrenectomy, which revealed renal cell carcinoma in the kidney and non-Hodgkin's B-cell lymphoma of the diffuse large cell type in the adrenal grand. The patient has been disease-free for ten months after the operation. To our knowledge, this is the first report of coexisting renal cell carcinoma and adrenal lymphoma. The relationship between these two disorders in our patient is discussed.

Ionotropic and metabotropic GABA and glutamate receptors in primate basal ganglia.

The functions of glutamate and GABA in the CNS are mediated by ionotropic and metabotropic, G protein-coupled, receptors. Both receptor families are widely expressed in basal ganglia structures in primates and nonprimates. The recent development of highly specific antibodies and/or cDNA probes allowed the better characterization of the cellular localization of various GABA and glutamate receptor subtypes in the primate basal ganglia. Furthermore, the use of high resolution immunogold techniques at the electron microscopic level led to major breakthroughs in our understanding of the subsynaptic and subcellular localization of these receptors in primates. In this review, we will provide a detailed account of the current knowledge of the localization of these receptors in the basal ganglia of humans and monkeys.

Defective morphogenesis and functional maturation in fetal islet-like cell clusters from OLETF rat, a model of NIDDM.

A failure in the compensate proliferation of pancreatic beta-cells, as the primary pathogenic event, has been reported in OLETF rat, a model of NIDDM. The aim of the present study is to define whether the beta-cell defect is attributed to the fetal stage islet development, if so, whether the defect involves down regulation of PDX-1 protein expression. Morphological changes, beta-cell function, and the expression of PDX-1 protein were examined in the cultured fetal islet-like cell clusters (ICCs) from OLETF rats along with their diabetes-resistant control counterpart LETO rats in the presence of 5.5 or 11.1mM glucose for 48, 72, 96, and 120-hr, respectively. We have observed four abnormalities in the ICCs of OLETF rats. First, a defective morphogenesis was noted during the 72 to 120-hr ICC culture, a period characterized by a dramatic increase in both beta-cell and non-beta-cell (alpha, delta, and PP) populations in control rats. This defective morphogenesis was demonstrated by a growth retardation of epithelial stratification and poor development of both beta-cell and non-beta-cell masses along with a parallel decline in relevant islet hormone contents. Second, a functional defect was characterized by failure to response to glucose during the 96 to 120-hr-cultured ICCs. Third, the ultrastructural analysis revealed a significant reduction in the number of secretory granules. Four, Western blot analysis showed a significant decrease of PDX-1 protein expression in the OLETF ICCs cultured in 11.1mM glucose for 48 to 72-hr and in 5.5mM glucose for 120-hr. Therefore, we concluded that during the fetal stage of islet development, OLETF rats exhibit both morphological and functional defects.

[Angioimmunoblastic T-cell lymphoma presenting with rapidly increasing biclonal gammopathy].

A 59-year-old man was admitted to our hospital with high fever and dysphagia. Physical examination revealed a lingual tumor and swelling of the cervical and inguinal lymph nodes. A lymph node biopsy specimen revealed that the normal nodal architecture had been obliterated by infiltration of small, intermediate, and large pale lymphocytes. Arborous branching of high endothelial venules was also observed. Southern blotting analysis showed rearrangement of the T-cell receptor beta gene. A diagnosis of angioimmunoblastic T-cell lymphoma was made. Serial serum protein fractionation analysis showed rapidly increasing M-protein. Immunofixation electrophoresis indicated specific bands for IgG kappa and IgM lambda, confirming the presence of biclonal gammopathy. Biclonal gammopathy has rarely been reported in patients with angioimmunoblastic T-cell lymphoma. The relationship between the occurrence of biclonal gammopathy and the possible role of HHV-6, HHV-8 and EBV is discussed.

[Introduction and some problems of the rapid time series laboratory reporting system].

We introduced an on-line system of biochemical, hematological, serological, urinary, bacteriological, and emergency examinations and associated office work using a client server system NEC PC-LACS based on a system consisting of concentration of outpatient blood collection, concentration of outpatient reception, and outpatient examination by reservation. Using this on-line system, results of 71 items in chemical serological, hematological, and urinary examinations are rapidly reported within 1 hour. Since the ordering system at our hospital has not been completed yet, we constructed a rapid time series reporting system in which time series data obtained on 5 serial occasions are printed on 2 sheets of A4 paper at the time of the final report. In each consultation room of the medical outpatient clinic, at the neuromedical outpatient clinic, and at the kidney center where examinations are frequently performed, terminal equipment and a printer for inquiry were established for real-time output of time series reports. Results are reported by FAX to the other outpatient clinics and wards, and subsequently, time series reports are output at the clinical laboratory department. This system allowed rapid examination, especially preconsultation examination. This system was also useful for reducing office work and effectively utilize examination data.

Behavior-related changes in the activity of substantia nigra pars reticulata neurons in freely moving rats.

As one of the primary targets of the striatum, the substantia nigra pars reticulata (SNr) has been hypothesized to play a role in normal motor behavior. Specifically, inhibition of usually high, tonic SNr output is predicted to correlate with motor activation. While support for this has come primarily from electrophysiological studies in primates performing goal-directed movements, we tested this hypothesis in rats behaving in an open-field arena. SNr single-unit activity was recorded during spontaneous bouts of open-field behavior (e.g., head and body movements, locomotion) and after rats were given D-amphetamine (1.0 mg/kg, s.c.), which reliably increases motor activity and elevates the firing of motor-related striatal neurons. Prior to drug administration, SNr neurons had either regular, slightly irregular or irregular firing patterns when animals rested quietly. During movement, some inhibitions were observed, but the majority ( approximately 79%) of analyzed units increased firing by as much as 38%. Regardless of the predrug behavioral response of the cell, amphetamine strongly inhibited firing rate ( approximately 90% below nonmovement baseline) and changed firing pattern such that all cells fired irregularly. Subsequent injection with the dopamine antagonist haloperidol (1.0 mg/kg, s.c.) reversed amphetamine-induced inhibitions in all tested cells, which supports a role for dopamine in this effect. These results suggest that the pattern of striatal activity established by amphetamine, which may be critical for determining the drug-induced behavioral pattern, is represented in the SNr regardless of the predrug behavioral response of the cell.

Immunological abnormalities in islets at diagnosis paralleled further deterioration of glycaemic control in patients with recent-onset Type I (insulin-dependent) diabetes mellitus.

AIMS/HYPOTHESIS: To determine whether the clinical heterogeneity observed in the development of Type I (insulin-dependent) diabetes mellitus correlates with immunohistochemical differences observed at diagnosis. METHODS: Patients (n = 17) with recent-onset diabetes clinically considered to be insulin dependent (Type I), underwent pancreatic biopsy for immunohistological analysis. These patients were divided into two groups based on the presence or absence of islet immunological abnormalities (insulitis or hyperexpression of MHC class I antigens or both). The patients were also HLA typed and tested for islet cell antibodies and antibodies to glutamic acid decarboxylase (GAD-Ab). All patients were followed monthly for 2 years and their fasting plasma glucose, haemoglobin A1c and daily insulin doses were recorded. The clinical course of patients with islet immunological abnormalities was compared with that of patients without those abnormalities. RESULTS: Patients with and without islet immunological abnormalities did not differ with regard to HLA type or islet cell antibodies. Antibodies to glutamic acid decarboxylase correlated with the presence of insulitis and MHC class I hyperexpression. These local immunological abnormalities were also associated with higher haemoglobin A1c values (p < 0.05) and a trend towards greater insulin requirements. Further, patients with the islet abnormalities had higher fasting plasma glucose concentrations 2 years after the biopsy than at the time of the biopsy (p < 0.05). CONCLUSION/INTERPRETATION: The heterogeneous clinical course observed following diagnosis in patients with Type I diabetes correlates with islet immunological abnormalities. Insulitis and hyperexpression of MHC class I correlate with deteriorating glycaemic control.

Changes in islet capillary angioarchitecture coincide with impaired B-cell function but not with insulin resistance in male Otsuka-Long-Evans-Tokushima fatty rats: dimorphism of the diabetic phenotype at an advanced age.

The Otsuka-Long-Evans-Tokushima fatty (OLETF) rat is a genetic model of spontaneous development of non-insulin-dependent diabetes mellitus (NIDDM) established as an inbred strain after 20 generations of selective breeding. Although they are thought to be genetically homogeneous, they show a dimorphism regarding the diabetic phenotype at an advanced age, with one remaining obese and modestly diabetic while the other becomes lean and overtly diabetic. To clarify the causes for this divergence, we examined the physical, biochemical, and histopathological features in rats at 50 weeks of age, including an analysis of islet angioarchitecture. Sixty-one of 85 male OLETF rats lost weight, while the remainder remained obese. Mean nonfasting plasma glucose in the lean group was 21.8+/-4.6 mmol/L, significantly higher versus the obese group (10.5+/-1.4 mmol/L) and the age-matched control Long-Evans-Tokushima-Otsuka (LETO) group (7.1+/-0.6 mmol/L). Morphological studies of the pancreas from the lean group showed enlarged multilobulated fibrotic islets with a paucity of B cells, whereas islets from the obese group appeared slightly enlarged and showed a relative abundance of B cells. The fine capillaries that form a network in the islets were extremely sparse in the lean group, resulting in a defective glomerular-like configuration, whereas those from the obese group were dense, forming a nearly typical glomerular-like configuration. Increased plasma insulin responses to oral and intravenous (i.v.) glucose and i.v. glucagon loads were nearly absent in the lean group, while they were evident in the obese group, although to a lesser extent compared with the LETO group. Mean insulin secretory output from the perfused pancreas in response to 11.1 mmol/L glucose in the lean group (3.5+/-2.2 pmol/20 min) was significantly lower versus the obese group (8.8+/-6.5 pmol/20 min) and LETO group (22.0+/-10.8 pmol/20 min). Similarly, pancreatic insulin content was significantly lower in the lean group (9.3+/-6.1 microg) versus the others (26.1+/-17.3 microg for obese and 41.1+/-24.8 microg for LETO). In vivo insulin-stimulated glucose uptake measured by a euglycemic clamp technique was significantly higher in the lean group compared with the obese group. These results demonstrate that the dimorphism regarding the diabetic phenotype in male OLETF rats at 50 weeks of age was due to differences in the number of islet B cells, which could be the result of a variation in the capacity for B-cell proliferation among male OLETF rats.

Dysfunctions of the epididymis as a result of primary carnitine deficiency in juvenile visceral steatosis mice.

The juvenile visceral steatosis mutant mice serve as an animal model of primary carnitine deficiency, classified as the sudden infant death syndrome. The defect in carnitine uptake was recently found to be due to a defect in the carnitine transporter gene. We herein report, for the first time, the characteristics of epididymal dysfunction in juvenile visceral steatosis mice. At 8-9 weeks of age, the epididymis was deformed and weight was significantly increased. Histologically, the duct of the proximal epididymis was dilated due to the accumulation of an unusually high level of spermatozoa. Spermatozoa were extravasated from the epididymal duct into the stroma. In contrast, the duct of the distal epididymis was constricted and contained no spermatozoa. Thus, the epididymal disorder causes obstructive azoospermia, leading to infertility.

Pharmacokinetic analysis of the cardioprotective effect of 3-(2,2, 2-trimethylhydrazinium) propionate in mice: inhibition of carnitine transport in kidney.

The site of action of 3-(2,2,2-trimethylhydrazinium) propionate (THP), a new cardioprotective agent, was investigated in mice and rats. I.p. administration of THP decreased the concentrations of free carnitine and long-chain acylcarnitine in heart tissue. In isolated myocytes, THP inhibited free carnitine transport with a Ki of 1340 microM, which is considerably higher than the observed serum concentration of THP. The major cause of the decreased free carnitine concentration in heart was found to be the decreased serum concentration of free carnitine that resulted from the increased renal clearance of carnitine by THP. The estimated Ki of THP for inhibiting the reabsorption of free carnitine in kidneys was 52.2 microM, which is consistent with the serum THP concentration range. No inhibition of THP on the carnitine palmitoyltransferase activity in isolated mitochondrial fractions was observed. These results indicate that the principal site of action of THP as a cardioprotective agent is the carnitine transport carrier in the kidney, but not the carrier in the heart.

Cardiomegaly in the juvenile visceral steatosis (JVS) mouse is reduced with acute elevation of heart short-chain acyl-carnitine level after L-carnitine injection.

The long-term administration of L-carnitine was very effective in preventing cardiomegaly in juvenile visceral steatosis (JVS) mice, which was confirmed by heart weight as well as the lipid contents in heart tissue. After i.p. injection of L-carnitine, the concentration of free carnitine in heart remained constant, although serum free carnitine level increased up to 80-fold. On the other hand, a significant increase in short-chain acyl-carnitine level in heart was observed. These results suggest that increased levels of short-chain acyl-carnitine, not free carnitine, might be a key compound in the protective effect of L-carnitine administration in JVS mice.

Aromatase inhibition reduces dendritic growth in a sexually dimorphic rat spinal nucleus.

The rat lumbar spinal cord contains the steroid-sensitive spinal nucleus of the bulbocavernosus (SNB), whose motoneurons innervate perineal muscles involved in copulatory reflexes. In normal males, SNB motoneuron dendrites grow exuberantly through postnatal (P) day 28. This growth is steroid dependent: Dendrites fail to grow in males castrated at P7, but grow normally in castrates treated with testosterone or its metabolites, dihydrotestosterone combined with estrogen. Treatment with either metabolite alone supports dendritic growth, but not to the level of testosterone-treated or intact males. In this study, we tested the hypothesis that aromatization of androgens to estrogens was involved in the masculine development of SNB dendrites. Motoneuron morphology was assessed in normal males and males treated daily (P7-28) with fadrozole, a potent aromatase inhibitor (0.25 mg/kg, subcutaneously) or saline vehicle (n = 4-6/group). SNB motoneurons were retrogradely labeled with cholera toxin-horseradish peroxidase at P28 (when dendritic length is normally maximal) and reconstructed in three dimensions. Comparable labeling was seen across groups; it was equivalent in both the rostrocaudal and radial extents. However, dendritic lengths in fadrozole-treated males were significantly below those of intact or saline-treated males. Neither SNB somata size nor target muscle weight differed across groups. These results suggest that aromatization of androgens to estrogens is necessary for development of masculine SNB dendritic morphology.

In vivo effects of calcium entry blockers on human parathyroid adenoma cells with special reference to calcium sensing ability and the hormone secretion.

We evaluated the effects of calcium-entry blockers on parathyroid hormone (PTH) secretion by human parathyroid adenoma cells in vitro. Nifedipine and bamidipine inhibited PTH secretion, while diltiazem had no significant effect. Cytosolic calcium concentrations were measured by use of the calcium-sensitive fluorescent dye fluo-3 with confocal laser scanning microscopy. Nifedipine increased the cytosolic concentration of calcium, whereas diltiazem decreased it. Results suggest that, in parathyroid adenoma cells, regulation of PTH secretion with respect to intracellular calcium concentration would be maintained despite differing response of intracellular calcium concentration following exposure to calcium-entry blockers.

Immunohistochemical localization of betacellulin, a new member of the EGF family, in normal human pancreas and islet tumor cells.

Betacellulin (BTC) purified from mouse beta cell tumor (betaTC-3) is a new member of the epidermal growth factor (EGF) family which can bind receptor tyrosine kinase, EGF receptor (erbB1) and erbB4. It has been demonstrated that proBTC mRNA was abundantly expressed in human pancreas tissue, and that BTC converted amylase-secreting rat acinar cell line (AR42J) into insulin-secreting cells, suggesting that BTC might be important for the growth and/or differentiation of islet cells. However, the cell type producing BTC in the pancreas has not been clarified. In this study, we examined the localization of BTC in human pancreas and islet cell tumors. Immunohistochemistry using specific antibodies to human BTC revealed that this protein was produced in alpha cells and duct cells, and probably in beta cells in normal adult pancreas. Furthermore, strong immunoreactivity to BTC was detected in primitive duct cells of the fetal pancreas, and both insulinoma and glucagonoma cells also showed positive immunoreactivity to BTC. EGF receptor (erbB1) and erbB4 were expressed mainly in islet and duct cells, and duct cells, respectively. These results demonstrate the localization of BTC and its receptors, and suggest that BTC may be one of the factors that have physiologically important roles such as growth and differentiation of islet cells in the human pancreas.

A missense mutation of mouse OCTN2, a sodium-dependent carnitine cotransporter, in the juvenile visceral steatosis mouse.

Carnitine is an essential cofactor for the mitochondrial beta-oxidation of long-chain fatty acids. The juvenile visceral steatosis (JVS) mouse, an animal model of systemic carnitine deficiency, is inherited in an autosomal recessive manner. Recently, a human OCTN2 gene encoding a sodium-dependent carnitine cotransporter was isolated and mapped to human chromosome 5q31. Since the mouse jvs locus was assigned to the region of chromosome 11 where it is syntenic to human chromosome 5q31, we isolated the mouse octn2 gene and screened for its mutation in the jvs mouse. DNA sequencing analysis disclosed a missense mutation from CTG (Leu) to CGG (Arg) at codon 352 located within the sixth transmembrane domain of octn2. This amino acid replacement possibly causes the conformational change of the protein that leads to dysfunction of the gene product. Hence, we conclude that octn2 is a candidate gene responsible for the JVS mouse.