LFA-1 and ICAM-1 molecule expression in jejunal mucosa from children with autoimmune enteropathy.

The expression of adhesion molecules by cells of the small intestinal mucosa was compared in gut biopsies from children with autoimmune small intestinal enteropathy and normal controls and related to HLA-DR expression by the same tissue. Jejunal biopsies were stained by IFL with monoclonal antibodies to LFA-1 (TS1/22 and CD11a/25.3.1) and ICAM-1 (RR1/1 and 84H10) molecules. LFA-1 and ICAM-1 positive cells were observed in the lamina propria in all cases and the counts were increased in autoimmune enteropathy compared with controls. In addition, in 4 of 7 cases of autoimmune enteropathy crypt enterocytes were positives for ICAM-1 when stained with RR1/1 and 3 of the 4 were also positive for LFA-1 when stained with both LFA-1 reagents. We speculate on the role of adhesion molecule expression in autoimmune enteropathy.


Analysis of heart rate variability to assess hemodynamic alterations following induction of anesthesia.

Extensive changes in hemodynamics and cardiac rhythm during induction of anesthesia may be mediated by altered responses of the autonomic nervous system to anesthetic agents. Analysis of the power spectrum of the heart rate (PSHR) variability can supply information about the autonomic nervous system, and may be used in order to assess this phenomenon. In this study, 78 patients undergoing coronary artery bypass graft surgery were evaluated. Anesthesia was induced with sufentanil, and neuromuscular blockade with vecuronium, a combination that may cause a decrease in heart rate. Before and after induction of anesthesia, the heart rate (HR), blood pressure (BP), cardiac output (CO), cardiac index (CI), and PSHR components were recorded. PSHR was obtained by using a special algorithm and data acquisition system for real-time spectral analysis. A low-frequency component (LFa, mainly sympathetic) was analyzed from a band of 0.04 Hz to 0.1 Hz. A high-frequency component (RFa, parasympathetic) was identified by the respiratory frequency spectrum. Alterations of the heart rate after induction of anesthesia were defined in order to separate the patient population into two groups: slow heart rate (slow-HR) and stable heart rate (stable-HR). Slow heart rate was defined as a decrease in HR of more than 20% of the baseline value. The variables were analyzed and compared between the slow-HR (n = 25) and stable-HR (n = 53) groups in order to verify the possibility of identifying patients prone to hemodynamic changes after anesthesia induction. There were no differences in preoperative HR, BP, CO, or CI between groups before anesthesia induction.(ABSTRACT TRUNCATED AT 250 WORDS)


Differential diagnosis of HTLV-I-associated myelopathy and multiple sclerosis in Iranian patients.

Two Iranian patients with chronic progressive spastic paraparesis and urinary dysfunction were referred to our hospital with the presumptive diagnosis of multiple sclerosis (MS). Routine CSF analysis and magnetic resonance imaging of the two patients were only partially characteristic of MS. Testing for antibodies to human T-cell leukemia virus type I [HTLV-I] in serum using a radioimmune precipitation assay revealed antibodies to HTLV-I in both patients. The infection with HTLV-I was confirmed by polymerase chain reaction (PCR) and liquid hybridization analysis using primers to the tax/rex region and a corresponding probe, demonstrating proviral DNA in peripheral blood mononuclear cells of both patients. On the basis of these findings demonstrating the presence of proviral HTLV-I DNA in the two Iranian patients, the initial diagnosis of MS was corrected to that of HTLV-I-associated myelopathy (HAM). In contrast, several patients with definite MS (nine from Germany, two from Iran) with a relapsing and remitting form of the disease were tested for HTLV-I infection by enzyme-linked immunosorbent assay and PCR, which yielded negative results. However, the mother of one HAM patient was found to be infected with HTLV-I. To support an association between HTLV-I infection and CNS disease in the two HAM patients, we analyzed the production of specific IgG antibodies within the CNS based on a simple enzyme immunoassay for viral IgG antibodies in CSF and serum. In the two HAM patients there was significant intrathecal antibody production directed against HTLV-I, but this was not found in any of the samples from MS patients.(ABSTRACT TRUNCATED AT 250 WORDS)


Balanced efficiencies of splicing and cleavage-polyadenylation are required for mu-s and mu-m mRNA regulation.

The relative abundance of the RNAs encoding the membrane (mu-m) and secreted (mu-s) forms of immunoglobulin mu heavy chain is regulated during B cell maturation by a change in the mode of RNA processing. This regulation depends on a competition between two mutually exclusive RNA processing reactions, cleavage-polyadenylation at the microseconds poly(A) site and splicing of the Cmu4 and M1 exons. Previously, the efficiencies of these two reactions were altered independently. When an efficient processing signal replaced the normal suboptimal signals of the mu gene, a single RNA product was produced exclusively. In this report, two efficient signals are combined in a single mu transcript and shown to restore a processing balance such that two mRNAs can once again be alternatively processed from a single RNA precursor. The ratio of the two RNAs generated from these mu genes containing balanced competing strong splice and cleavage-polyadenylation reactions display the expected developmental shift when expressed in B cells and plasma cells. Therefore, the balance between cleavage-polyadenylation and splicing efficiencies is critical to the developmentally regulated expression of mu-s and mu-m mRNA. Also shown here is that the entire mu-m region, including the M1 and M2 exons and the mu-m poly(A) site, can be replaced with SV40 splice and poly(A) sequences. Regulation is maintained in these mu genes, indicating that no specific sequences within the mu-m region are required.


Occupancy-response relationships for beta- and alpha 2-adrenergic receptors exerting opposing effects on cAMP production.

In primary glial cultures, norepinephrine (NE) activates both beta-adrenergic receptors to increase cAMP formation and alpha 2-adrenergic receptors to partially inhibit this response. We used selective alkylating agents to compare the concentration-dependence and receptor reserves for activation of each subtype. Partial inactivation of beta-receptors with alkylating pindolol (BIM) caused a slight decrease in the potency of isoproterenol (ISO) in increasing cAMP accumulation and a progressive decrease in maximum response. The KA for ISO was 9.8 +/- 2 nM, with a 2-3-fold beta-receptor reserve. BIM pretreatment decreased the maximal response to NE without significantly altering its apparent EC50 (41 +/- 6.7 nM). Partial inactivation of alpha 2-adrenergic receptors with EEDQ increased the maximal response to NE without significantly altering its apparent EC50 (41 +/- 6.2 nM). NE inhibited the cAMP response to ISO with an apparent EC50 of 38 +/- 1.2 nM. EEDQ pretreatment reduced inhibition of the ISO response by both NE and the alpha 2-agonist UK 14,304, and inhibition of the forskolin response by UK 14,304. EEDQ pretreatment caused only a small decrease in potency for the alpha 2-agonists. The KA for NE in inhibiting the ISO response was 120 +/- 30 nM, indicating a 2-3-fold alpha 2-receptor reserve. These results suggest that NE has similar affinities and receptor reserves for beta- and alpha 2-adrenergic receptors in this system, and activates and inhibits adenylate cyclase at the same agonist concentrations.


Human alpha-calcitonin gene-related peptide stimulates adenylate cyclase and guanylate cyclase and relaxes rat thoracic aorta by releasing nitric oxide.

1. The signal transduction pathway for vasorelaxation induced by human alpha-calcitonin gene-related peptide (human alpha-CGRP) was studied in rat thoracic aortic rings preconstricted with noradrenaline (10(-7) M). 2. Vasorelaxation by human alpha-CGRP was inhibited by haemoglobin (10(-6) M) and methylene blue (10(-5) M) but was unaffected by ibuprofen (10(-5) M). 3. Acetylcholine caused a 16 fold increase in levels of guanosine 3':5'-cyclic monophosphate (cyclic GMP) with levels of adenosine 3':5'-cyclic monophosphate (cyclic AMP) being unaltered. Human alpha-CGRP caused a 12 fold increase in levels of cyclic GMP but, in contrast to acetylcholine, evoked a 2.5 fold rise in levels of cyclic AMP. The rises in cyclic nucleotides evoked by human alpha-CGRP and acetylcholine were dependent on the presence of an intact endothelium. 4. NG-nitro-L-arginine (L-NOARG: 10(-5) M), which inhibits nitric oxide synthetase, inhibited the relaxant response to human alpha-CGRP and cyclic GMP accumulation without affecting the cyclic AMP accumulation. 5. The data presented in this paper suggests that human alpha-CGRP relaxes the rat thoracic aorta by releasing nitric oxide and stimulating guanylate cyclase. The stimulation of adenylate cyclase by human alpha-CGRP probably precedes the activation of nitric oxide synthase but could be unrelated to the relaxant response.


Identification of alpha 1-adrenoceptor subtypes in the rat vas deferens: binding and functional studies.

1. The alpha 1-adrenoceptor subtypes of the prostatic and epididymal portion of rat vas deferens were characterized in binding and functional experiments. 2. In saturation experiments, [3H]-prazosin bound to two distinct affinity sites in the epididymal portion of rat vas deferens (pKD = 10.1 +/- 0.13 and 9.01 +/- 0.15, Bmax = 507 and 1231 fmol mg-1 protein, respectively). In the prostatic portion [3H]-prazosin bound to a single affinity site (pKD = 9.82 +/- 0.04, Bmax = 924 fmol mg-1 protein). 3. In the displacement experiments, unlabelled prazosin displaced biphasically the binding of 200 pM [3H]-prazosin to the epididymal portion; the resulting two pKI values were consistent with the affinity constants obtained in the saturation experiments. WB4101 (2-(2,6-dimethoxy-phenoxyethyl)-amino-methyl-1,4-benzodioxane) and benoxathian also discriminated the two affinity sites in the epididymal portion and the population of low affinity sites for the three antagonists was approximately 40%. On the other hand, the prostatic portion predominantly showed a single affinity site for prazosin, WB4101 and benoxathian, although the presence of a small proportion (less than 10%) of the low affinity site could be detected. HV723 (alpha-ethyl-3,4,5-trimethoxy-alpha-(3-((2-(2-methoxyphenoxy)ethyl)-a min o)- propyl) benzeneacetonitrile fumarate) displaced the [3H]-prazosin binding monophasically with a low affinity in both halves. 4. Pretreatment with chlorethylclonidine (CEC) at concentrations higher than 1 microM inhibited 700 pM [3H]-prazosin binding to the prostatic portion by approximately 50%. However, the inhibition in the epididymal portion was much less (approximately 21% at 50 microM CEC).5. In the functional study, the contractile response to noradrenaline was competitively inhibited by prazosin, WB4101, benoxathian and HV723 with similar and low affinities (pKB value ranging from 8.0to 9.0) in the epididymal portion of rat vas deferens. In the prostatic portion of rat vas deferens,noradrenaline also produced a contraction, but the maximal amplitude of contraction developed was approximately one-fourth of that in the epididymal portion. Prazosin and WB4101 also inhibited the contractile response of the prostatic portion with the pKB values similar to those obtained in the epididymal portion. The contractions to noradrenaline in both portions were potently attenuated by 1 LM nifedipine but were not affected by pretreatment with 1O LM CEC.6. Under conditions where P2x-purinoceptors and prejunctional M2-adrenoceptors were blocked, electrical transmural stimulation produced a rapidly developing phasic contraction and a subsequent tonic contraction in the epididymal portion of rat vas deferens. The phasic and tonic contractions were inhibited in a concentration-dependent manner by prazosin (ICs = 25.7 and 25.9 nm, respectively),WB4101 (ICo= 7.27 and 7.58 nM), benoxathian (ICs = 10.9 and 8.66 nM) and HV723 (ICs = 15.9 and 14.9 nM). Nifedipine selectively attenuated the tonic contraction induced by electrical stimulation, and the residual phasic response was inhibited by the antagonists mentioned above with similar affinities to those in the absence of nifedipine. CEC (10 gM) had little effect on the adrenergic neurogenic contractions.7. The present results indicate the presence of two distinct alpha&-adrenoceptor subtypes in the rat vas deferens, which show respectively high and low affinities for each of prazosin, WB4101 and benoxathian,and presumably correspond to putative MIA and alL subtypes according to the recent am-adrenoceptorsubclassifications. The contractions induced by exogenous and endogenous noradrenaline seem to be predominantly mediated through the alL subtype. The heterogeneous distribution of the low affinity sites(alL subtype) may well explain differences in functional responsiveness between the two portions of rat vas deferens.


Multiple sigma binding sites in guinea-pig and rat brain membranes: G-protein interactions.

1. Evidence is accumulating for multiple sigma (sigma) sites in the mammalian CNS. 2. We have addressed this problem and have examined sigma site - G-protein coupling in guinea-pig and rat brain membranes. 3. Ditolylorthoguanidine (DTG), (+)-3-(3-hydroxyphenyl)-N-1-(propyl)piperidine (3PPP) and dextromethorphan displaced [3H]-DTG (3.4 nM) with low Hill slopes of 0.5, 0.6 and 0.6, respectively in guinea-pig brain membranes. 4. In the presence of 5'-guanylylimidodiphosphate (Gpp(NH)p; 100 microM), the specific binding of [3H]-DTG was reduced by 36.7%, the Hill slope of 3PPP was increased to near unity, the ability of dextromethorphan to displace DTG was virtually abolished and the Hill slope for DTG remained low (0.7), indicating the presence of at least two binding sites. These data indicate that although Gpp(NH)p removes a dextromethorphan high affinity site, two DTG selective sites remain in the presence of Gpp(NH)p. 5. The present study suggests that DTG binds to at least three sites in guinea-pig brain membranes, at least one of which is G-protein linked. 6. In rat brain membranes, DTG displaced itself (3.4 nM) with a Hill slope near 1. 3PPP displacement of [3H]-DTG was comparable with the guinea-pig (Hill slope 0.5) and displaced from more than 1 site. Dextromethorphan did not displace [3H]-DTG at concentrations below 10 microM. 7. The heterogeneity of sigma sites appears to be less in rat than in guinea-pig brain membranes.


Muscarinic blockade of beta-adrenoceptor-stimulated adenylyl cyclase: the role of stimulatory and inhibitory guanine-nucleotide binding regulatory proteins (Gs and Gi).

1. The functional antagonism that exists between muscarinic and beta-adrenoceptor function in guinea-pig tracheal smooth muscle was investigated by assessing Gs and Gi regulated adenylyl cyclase activity in isolated membranes. 2. Membranes from guinea-pig tracheal smooth muscle contain both Gi alpha and Gs alpha as assessed by Western blots with anti-G-protein antibodies. 3. GppNHp, a non-hydrolysable analogue of guanosine 5'-triphosphate (GTP), was shown to stimulate adenylyl cyclase activity at high concentrations (10(-6)-10(-4) M). GppNHp also produced a concentration-dependent reduction in pertussis toxin-catalysed adenosine diphosphate (ADP)-ribosylation of Gi alpha. 4. Pretreatment of tracheal smooth muscle slices with methacholine (10(-6) M) provoked a blockade of isoprenaline plus GTP, GppNHp- and GTP-stimulated adenylyl cyclase. 5. Addition of methacholine to membranes did not trigger inhibition of GTP-stimulated adenylyl cyclase activity but did block the isoprenaline-mediated augmentation of GTP-stimulated adenylyl cyclase activity. 6. Pretreatment of tracheal smooth muscle with methacholine (10(-6) M) provoked a blockade of cholera toxin-catalysed NAD(+)-dependent ADP-ribosylation of Gs alpha. 7. Phorbol-12-myristate 13-acetate (PMA)-treatment of tracheal smooth muscle slices actually enhanced GppNHp-stimulated adenylyl cyclase activity in subsequently prepared membranes. 8. We suggest that methacholine in addition to inhibiting adenylyl cyclase via a Gi-dependent mechanism induces a functional inactivation of Gs activity. These results together may explain the functional antagonism that exists between increased muscarinic tone and the ability of beta-adrenoceptor agonists to provoke excitation-contraction uncoupling.


The glutamate antagonist, MK-801, does not prevent dopaminergic cell death induced by the 1-methyl-4-phenylpyridinium ion (MPP+) in rat dissociated mesencephalic cultures.

The neuroprotective effects of MK-801, a non-competitive antagonist of the N-methyl-D-aspartate (NMDA) receptor/channel, were assessed in a culture model which reproduces in vitro the selective degeneration of mesencephalic dopaminergic neurons seen in parkinsonian brains. Dissociated mesencephalic cells derived from rat embryonic brains were subjected for 24 h to intoxication by the 1-methyl-4-phenylpyridinium (MPP+), the active metabolite of the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MPP+ at 3 and 10 microM produced selective and dose-dependent damages to dopaminergic neurons as quantified by the loss of the number of TH immunoreactive cells and the loss of [3H]DA uptake whereas other cell types remained unaffected. MK-801 at 3 and 10 microM failed to rescue degenerating dopaminergic neurons in presence of MPP+. At 50 microM, i.e. the highest concentration that is not toxic by itself in this culture system, MK-801 was also found ineffective. Furthermore, degree of dopaminergic cell damage was not reduced when repeated additions of the glutamate antagonist (10 microM/6 h for 24 h) were performed during exposure to MPP+ or when mesencephalic cultures were left after intoxication for up to 2 days in a culture medium still supplemented with MK-801 but free of toxin. In accordance with these results, MK-801 did not affect significantly the uptake of [3H]DA in control cultures, thereby suggesting that this compound cannot prevent intracellular accumulation of MPP+ within dopaminergic neurons. At higher concentrations of MPP+ (100 microM) tested, toxic effects were seen toward dopaminergic neurons and non-dopaminergic cells as quantified by Trypan blue dye accumulation and loss of [3H]GABA uptake.(ABSTRACT TRUNCATED AT 250 WORDS)


Somatostatin release in rat neocortex during gamma-hydroxybutyrate-provoked seizures: microdialysis combined with EEG recording.

Gamma-hydroxybutyrate (GHB) was intracortically applied in two doses (first 10 and then 20 mg/ml) to awake Wistar rats using microdialysis. Simultaneously, EEG and the release of somatostatin-like immunoreactivity (SLI) were measured from the frontal cortex. Intracerebrally administered GHB induced cortical epileptogenic spikes, but not high voltage spindles (HVS) as reported after systemic administration, and seizures with myoclonic jerks and contraversive head movements. Compared to the basal level, GHB (10 mg/ml) initially increased the release of SLI (p < 0.05). However, when the frequency of spikes and seizures rose rapidly (p < 0.001), SLI release decreased significantly (p < 0.001). Minimum release of SLI occurred when seizures were most frequent (during perfusion with 20 mg/ml GHB), while after removal of the drug it rose above the basal level (p < 0.05). According to these results, intracortically applied GHB increases the release of SLI in the surrounding tissue. However, further exposure of GHB leads to a manifestation of epileptic spikes and seizures, during which the release of SLI is significantly attenuated. This suggests that release of somatostatin is affected during epileptic phenomena induced also by intracortical GHB application.


Hippocampal long-term potentiation in nucleus basalis magnocellularis-lesioned rats.

The probability of hippocampal long-term potentiation induction in the mossy fiber CA3 and commissural/associational CA3 responses and the cortical levels of choline acetyltransferase (ChAT) activity were compared in right nucleus basalis magnocellularis (NBM)-lesioned rats. A 50% reduction in the right cortical ChAT activity was demonstrated 4 weeks after an ibotenic acid lesion of the NBM. No significative differences were found in the probability of LTP induction of right hippocampal slices in sham-operated rats from 10 to 40 days after the injection into the right NBM. On the contrary, a progressive and significative increase in the probability of LTP induction was shown in right hippocampal slices of NBM-lesioned rats from 10 to 40 days after the injection of ibotenic acid into the right NBM. The results demonstrated the appearance of a paradoxical increase of hippocampal synaptic plasticity when the cortical cholinergic biochemical alterations are still present. This finding might be responsible for a behavioural recovery, in NBM-lesioned rats.


Somatostatin and growth hormone-releasing factor release from Zucker rat hypothalamic tissue.

Plasma somatotropin (ST) levels are depressed in the genetically obese Zucker rat compared to those of their littermates. It is believed that this defect is associated with one or both of the hypothalamic neuropeptides that control ST release: growth hormone releasing factor (GRF) and somatostatin (SS). The mechanism by which SS and GRF neuropeptides are regulated remains uncertain. The objective of this study was to examine the effect of 2 deoxy-glucose (2DG), isoproterenol (ISO), tryptophan (TRP), and 5HT on SS and GRF release in hypothalamic tissue from lean and obese Zucker rats. An in vitro perifusion system was established to examine the release of SS and GRF from perifused hypothalami taken from 8- and 12-week-old Zucker rats under basal conditions and in response to 2DG, ISO, TRP, 5HT, and KCl administration. Hypothalami were perifused with Dulbecco's modified eagle's medium continuously at 37 degrees C for 5 h at a flow rate of 100 ml/min. ISO and 2DG significantly (p < 0.05) increased SS levels from the obese rat, but no effect was observed from the lean littermate. GRF was not affected by 2DG or ISO in either genotypes. TRP and 5HT failed to affect SS or GRF release in lean or obese Zucker rats. It is proposed that the obese Zucker rat is more sensitive to glucose deprivation and to beta-adrenergic stimulation of SS release than the lean littermate.


Benzodiazepine receptor binding of nonbenzodiazepines in vivo: alpidem, zolpidem and zopiclone.

Several classes of nonbenzodiazepine compounds, including imidazopyridines such as alpidem and zolpidem and cyclopyrrolones, e.g., zopiclone, have effects similar to benzodiazepines and may act at the benzodiazepine receptor in brain. We characterized the binding of these compounds to the benzodiazepine site in three brain regions using specific uptake of the high-affinity ligand [3H]Ro15-1788 (flumazenil). For alpidem, benzodiazepine binding was decreased in cortex and hippocampus with increasing drug dose. For zolpidem, receptor binding was reduced in cortex without a dose-response effect and no effect was observed on cerebellar binding. Zopiclone did not alter binding except for a decrease in binding at the lowest dose evaluated and an increase in binding above control at the highest dose. These data corroborate prior studies indicating that the imidazopyridines appear to act at the benzodiazepine receptor, but do not support receptor subtype selectivity of zolpidem. The limited effect of zopiclone except for increased binding at high doses is also consistent with prior studies suggesting that zopiclone acts at a site distinct from the benzodiazepine receptor.


Complement 4 gene deletion in patients with IgA nephropathy and Henoch-Schönlein nephritis.

The fourth component of complement (C4), especially B isotype, has been said to be deficient in the IgA nephropathy and Henoch-Schönlein nephritis. However, the association between these diseases and C4 deficiency was questioned recently, and the usual C4 allotyping method is unable to discriminate the C4 deficiency from the C4 duplication. So by combining the DNA restriction fragment length polymorphism with the usual C4 allotyping, we tried to determine whether the deficiency of C4 can be demonstrated in the DNA level. We found that the frequency of C4 gene deletion was increased, although the frequency of null phenotype was not different from the control. From these results we can say that C4 gene deletion is a genetic risk factor in these diseases, at least in the Japanese population.


Tumor-specific chemoimmunotherapy of murine fibrosarcoma using tumor-specific transplantation antigen, cyclophosphamide, and interleukin-2.

The anti-tumor effect of active specific chemoimmunotherapy, using butanol-extracted tumor-specific transplantation antigen (TSTA), cyclophosphamide (CY), and continuous intrasplenic infusion of interleukin-2 (IS-IL-2), was assessed in a C3H/HeJ murine methylcholanthrene (MCA)-induced fibrosarcoma model. Sole administration of TSTA induced tumor-specific, suppressor T cells in the spleens of mice bearing 3-day established tumors. Concomitant low-dose (20 mg/kg) CY treatment not only inhibited TSTA-mediated suppressor cell induction, but also evoked splenic lymphocytes of tumor-bearing mice to display tumor-specific cytotoxic activity. High-dose (200 mg/kg) CY abrogated the immunotherapeutic benefit. The immune effectors generated by TSTA plus CY bear the Thy 1, L3T4, Lyt 2 phenotype. Continuous IS-IL-2 infusion in combination with TSTA and CY induced tumor-specific Lyt 2+ cytolytic T cells, as well as the activation of L3T4+ cytostatic T cells. Thus, a triple regimen using TSTA, CY, and IS-IL-2 appears to augment CTL induction in tumor-bearing hosts undergoing stimulation of helper elements by TSTA and inhibition of suppressor cells by CY.


Ethinyl estradiol-induced cell proliferation in rat liver. Involvement of specific populations of hepatocytes.

Hepatocyte proliferation was analyzed in vivo during the time course of continuous administration to rats of the liver tumor promoter ethinyl estradiol (EE) at 10 p.p.m. in the diet. EE-induced acute liver hyperplasia was detected in male and female Sprague-Dawley rats as an increased mitotic index of hepatocytes after 2 days of treatment. 5'-Bromodeoxyuridine (BrdU) labeling showed that proliferating hepatocytes were randomly distributed throughout the hepatic lobule. Subsequently, and still during the first few days of continuous EE treatment, hepatocyte proliferation decreased to control levels, and a transient increase in the incidence of apoptosis in the liver was detected. Although consistent with the concept of liver growth regression after mitogen-induced hyperplasia, these results differ from others reported to date in that, in our experiments, the cessation of cell proliferation and the subsequent growth regression occurred without withdrawal of EE in our experiments. After returning to control levels, hepatocellular proliferation again increased between 3 and 6 months of chronic treatment and remained activated during the following months of continuous treatment, as seen by accumulative BrdU labeling. Proliferating hepatocytes were predominantly located in zone 2 of the hepatic lobule at this time, surrounding a periportal zone of vacuolated hepatocytes, which were also induced by the treatment. Moreover, hyperplasia of basophilic hepatocytes was also seen around some portal spaces. In another set of experiments, chronic EE-induced activation was characterized by flow cytometry on hepatocytes isolated from male Fischer rats. Ploidy analysis of hepatocyte cell suspensions showed that the normal polyploid pattern of hepatocytes was altered by EE, the proportion of diploid hepatocytes rising considerably. The results also showed that these diploid cells were the most susceptible hepatocyte population to EE-induced proliferation, as shown by a combination of BrdU labeling and cell sorting methods. In contrast to Sprague-Dawley rats, no vacuolated cells were found histologically in the livers of these animals and the proliferating hepatocytes were located adjacent to the portal areas. These results taken together support the existence of cell target populations in the liver responding to the effects of tumor promoters. The finding that a subpopulation of diploid hepatocytes was the liver cell class most susceptible to proliferation during chronic EE treatment may explain, at least in part, the behavior of EE as a tumor promoter in hepatocarcinogenesis.


Proto-oncogene activation in liver tumors of hepatocarcinogenesis-resistant strains of mice.

Activation of the ras family of oncogenes occurs frequently in liver tumors of the B6C3F1 mouse, a strain which is highly sensitive to hepatocarcinogenesis. Many other mouse strains are much more resistant to hepatocarcinogenesis; the aim of this study was to determine the frequency and pattern of oncogene activation in spontaneous and chemically induced liver tumors of three such strains, the C57BL/6J, the C57BL/6 x DBA/2 F1 hybrid (B6D2F1) and the C57BL/6 x Balb/c F1 hybrid (B6BCF1). The C57BL/6, DBA/2 and Balb/c strains are all relatively resistant to spontaneous hepatocarcinogenesis (1.5-3.6% of animals develop liver tumors in 2 years); with regard to chemically induced hepatocarcinogenesis the Balb/c is highly resistant, the C57BL/6 has low susceptibility and the DBA/2 has low to moderate susceptibility. The nude mouse tumorigenicity assay was used to search for activated oncogenes in 15 C57BL/6J liver tumors induced by a single neonatal dose of vinyl carbamate (VC, 0.15 mumol/g body weight). Three tumors contained H-ras genes activated by point mutations at codon 61 and one contained a non-ras oncogene. The polymerase chain reaction and allele-specific oligonucleotide hybridization were used to study H-ras mutations in spontaneous and VC-induced tumors from all three strains of mice. The frequency of H-ras codon 61 mutations in tumors induced by 0.15 mumol/g body weight VC in the C57BL/6J mouse (5/37) was similar to that in spontaneous tumors (2/9); surprisingly, tumors induced by a lower dose of VC (0.03 mumol/g body weight) had a higher frequency of H-ras mutations (12/28). The frequencies of H-ras activation detected in VC (0.03 mumol/g body weight)-induced tumors from the two F1 hybrids studied differed markedly. Only one VC-induced B6BCF1 tumor contained a mutated H-ras gene (1/10), whereas the majority of B6D2F1 tumors contained such mutations (23/33). Several spontaneous B6D2F1 liver tumors contained H-ras codon 61 mutations (6/15). Thus, H-ras activation frequency does not determine susceptibility to hepatocarcinogenesis in inbred mice and their F1 hybrids, since a relatively high frequency of H-ras mutations was observed in two resistant strains and a low frequency was found in the other strain.


Expression of cell adhesion molecules and catecholamine synthesizing enzymes in the developing rat adrenal gland.

Cell adhesion molecules play a major role in determining tissue architecture during histogenesis. This immunocytochemical study of the adrenal gland examines the embryonic and early postnatal cellular expression of two neural cell adhesion molecules, NCAM and L1, which are widely expressed in brain and have been found also to be expressed in the adult rat adrenal gland. In parallel, antibodies directed against two neuroendocrine cell markers, tyrosine hydroxylase and phenylethanolamine N-methyltransferase, were employed to verify the phenotypic nature of developing chromaffin cells in order to correlate cell adhesion molecule expression with the state of chromaffin cell differentiation. NCAM was found to be expressed by chromoblasts within extra-adrenal blastema (i.e. before their migration into the cortical primordium) at the 16th day of embryonic life. It continued to be expressed by all developing chromaffin cells after their infiltration into the developing adrenal gland at all ages. L1 was also expressed by chromoblasts in extra-adrenal sites, but was found only in a subpopulation of chromaffin cells within the cortical primordium from the 16th embryonic day onwards. Those chromoblasts which expressed L1 constituted relatively large compact cell clusters within the gland at this stage, while intra-adrenal chromaffin cells not expressing L1 were dispersed in small cell groups. L1 was also strongly expressed by nerve fibres (and their surrounding Schwann cells) which appeared to innervate cell groups as early as the 16th embryonic day. Both extra- and intra-adrenal chromoblasts expressed tyrosine hydroxylase, but the large L1-positive cell aggregates were less intensely immunoreactive for tyrosine hydroxylase than were cells in small groups. PNMT expression was restricted to L1-negative intra-adrenal chromoblasts present in small groups. Ultrastructural observations demonstrated that cells expressing L1 contained few secretory granules at the 18th embryonic day. It is concluded from these data that these chromoblasts are the precursors of the noradrenergic cells found in the mature gland. In addition, the arrangement of noradrenergic chromaffin cells in the form of homotypic cell groups throughout the course of histogenesis of the adrenal medulla is likely to be a direct consequence of the exclusive co-expression of both NCAM and L1 by this subpopulation of maturing chromaffin cells.


Cellular expression of somatostatin in MAM-induced microencephaly in the rat.

Methylazoxymethanol acetate (MAM) is a mitotic inhibitor that has been used to selectively destroy neuroblasts at specific times during gestation. The administration of MAM results in a dose-dependent microencephaly. Following MAM treatment at 15 days of gestation, we have noted an increase in the level of SS immunoreactivity in the neocortex, as determined by radioimmunoassay. Northern blot analysis for preproSS mRNA revealed an increase in MAM-treated cortex. The cellular distribution of SS has been determined using in situ hybridization and immunocytochemistry. There was a 30% increase in the density of SS-immunoreactive neurons in the cortex of the MAM-treated animals. These data suggest that SS neurons in the cortex are spared following MAM treatment at GD 15.


Ultrastructure of spared dopamine terminals in caudate-putamen nuclei of adult rats neonatally treated with intranigral 6-hydroxydopamine.

Residual dopamine terminals in the dorsal striatum, caudate-putamen nuclei (CPN), of adult rats neonatally lesioned with 6-hydroxydopamine (6-OHDA) sustain a relatively high level of dopamine release. We examined whether there were morphological differences in the spared dopamine terminals that might correlate with this increased efficacy. Postnatal male rat pups from 50 litters were pretreated with desmethylimipramine (DMI) to protect from non-specific monoamine damage, then given unilateral intranigral injections of 6-OHDA or vehicle. Coronal sections through the CPN and substantia nigra of the surviving adult animals from each litter were co-processed for immunoautoradiographic or immunoperoxidase localization of the catecholamine synthesizing enzyme, tyrosine hydroxylase (TH). Quantitative ultrastructural analysis established that in animals showing maximal (greater than 90%) depletions in immunoautoradiographic labeling for TH, the number of TH-labeled axons in the CPN ipsilateral to the 6-OHDA injections was reduced to one third of the number seen in the contralateral, unlesioned hemisphere, or the CPN from vehicle-injected animals. The ultrastructural features of residual terminals ipsilateral to 6-OHDA lesions were morphologically similar to those of the contralateral side or in vehicle-injected animals. However, in comparison with controls, these TH-labeled terminals had significantly larger mean cross-sectional diameters. When subdivided into groups according to size, there were significantly fewer small (0.0-0.1 micron 2) and more large (0.41-0.50 micron 2) TH-immunoreactive profiles in lesioned versus unlesioned CPN. The remaining TH-labeled terminals ipsilateral to the 6-OHDA lesions also appeared to be more often in direct contact with unlabeled soma and proximal dendrites as opposed to dendritic spines in the unlesioned CPN. These results suggest that the enhanced activity of dopamine neurons innervating the CPN after nigral 6-OHDA lesions may contribute to changes in size and target of their terminals. Alternatively, the observed large size of remaining dopamine terminals may reflect selective vulnerability of smaller axons to 6-OHDA toxicity.


Sensory and motor innervation of bird intrafusal muscle fibers.

1. Most bird muscle spindles are supplied by only one primary afferent. 2. Secondary afferents occur irregularly. 3. Sensory terminals are covered by a basal lamina and a collagenous sheath. 4. Two types of motor terminal are recognized which can be referred to specific types of intrafusal fiber. 5. The sensory and motor innervation of bird intrafusal fibers is less understood than that of mammalian intrafusal fibers.


Effects of propionate on the acid microclimate of hen (Gallus domesticus) colonic mucosa.

1. Short-chain fatty acid absorption in hen colon is protonated across the apical border coupled to an apical electrogenic proton pump. 2. The surface pH of the isolated colonic epithelium was 6.27 +/- 0.05, when incubated in Krebs-phosphate buffer pH 7.0. 3. Propionate 7 and 40 mmol/l in the incubation medium (pH 7.0) increased microclimate pH to 6.47 +/- 0.04 and 6.56 +/- 0.04. Inhibition of metabolic activity by potassium cyanide 1 mmol/l increased surface pH to 6.66 +/- 0.06. 4. The calculated concentration of propionic acid in the microclimate is near-linearly related to the propionate concentration. Thus, the acid microclimate is not responsible for the Michaelis-Menten like kinetics of propionate transport.


Lipid reserves and body composition in postreproductive anurans.

1. The mass of lipid extracted from 13 species of adult anurans collected immediately after reproduction differed between sexes. 2. Covariance analyses of lipid mass with body mass as the covariate indicated that females had significantly more lipid than males; the proportion of lipid as a percentage of body mass was correlated significantly with water content of males, but was not correlated significantly in females. 3. Heat of combustion of extracted lipid increased among species in relation to the timing of reproduction; species that bred later in the year had a greater energy content per gram of lipid. 4. Major changes in composition during growth and metamorphosis of bullfrogs (Rana catesbeiana), include increases in lipid and energy content per gram of tissue interrupted by a severe decline of lipid reserve during metamorphosis.


Tetanus responses under rapid bath solution change: electrotonic depolarization of transverse tubules may release Ca2+ from sarcoplasmic reticulum of Rana japonica skeletal muscle.

1. Single skeletal muscle fibers were transferred from a normal Ringer solution to Na+ ion free solution, and vice versa, and tetanus responses were recorded immediately after the transfer. 2. Fractional tetanus tension recorded immediately after the displacement from the Na+ ion free solution to normal Ringer solution was dependent on fiber diameter. 3. Diffusion of Na+ ions along the transverse tubules was simulated [apparent diffusion constant was 3.11 x 10(-6) (cm2/s)]. 4. Our results suggest that the electronic spreading of membrane potential, caused by an action potential in the transverse tubules, could release Ca2+ ions from sarcoplasmic reticulum.


Epidermal growth factor alters the electrolyte profile of lactating ewes (Ovis aries).

1. Lactating ewes were treated with mouse epidermal growth factor (EGF) at a dose rate of 0.5 mg/day for 4 days and its effects on the electrolyte profile were observed. 2. There was no effect of EGF on plasma concentrations of sodium or potassium, although urinary and total (in urine and milk) losses of both were reduced. 3. EGF-induced hypocalcaemia was associated with reduced milk calcium secretion and increased urinary calcium excretion whereas EGF-induced hypermagnesaemia was associated with reduced urinary and total magnesium losses. 4. Glomerular filtration rate was reduced during EGF infusion. 5. Chronic intravenous EGF infusion affects the electrolyte profile by altering electrolyte secretion by the mammary gland and renal electrolyte excretion.


Physiological responses to exercise and hypoglycaemia stress in pigs of differing adrenal responsiveness.

1. Twelve Large White x Landrace male pigs, six with high adrenocortical response to ACTH, and six with low response, were subjected to mild and moderate exercise, and then to insulin-induced hypoglycaemia. 2. Plasma ACTH, cortisol, catecholamines and some haematological and plasma biochemical parameters were determined in response to exercise, and glucose and cortisol in response to insulin challenge. 3. High responders had significantly greater increases than low responders in ACTH, cortisol and catecholamines following exercise, and in cortisol following insulin challenge. 4. The results suggest that differences in adrenocortical response to exogenous ACTH are an accurate reflection of the animal's response to stressful stimuli.


Factors affecting oxygen consumption in wild-caught yellow-bellied marmots (Marmota flaviventris).

1. All age groups gained mass during the active season, but mass-gain of adult females was delayed during lactation. 2. The relationship of body mass to metabolic rate varied widely; when the relationship was significant, R2 varied from 10.3 to 72.6%. Body mass affects VO2 more during lactation than at any other period. 3. Mean VO2 of adult males was higher in June than that of adult, non-lactating females. 4. VO2 of reproductive females was significantly higher during lactation than during gestation or postlactation because specific VO2 varied. Specific VO2 of non-reproductive females declined over the active season. 5. Specific VO2 of all age groups declined between the premolt and postmolt periods. The reduced maintenance costs can contribute 20-46% to daily growth. 6. Observed VO2 was lower than the value predicted from intraspecific or interspecific Bm:M regressions. 7. VO2 of wild-caught marmots was lower than that of marmots maintained in the laboratory, probably because of dietary differences. 8. Because basal metabolism is a stage on a food-deprivation curve, we suggest that basal metabolic rate is not an appropriate measure of the metabolic activity of free-ranging animals.


Specificity of the permissive effect of D-glucose on insulin release in chicken pancreas.

1. As previously shown, 14 mM D-glucose, a non-insulinotropic concentration in isolated chicken pancreas, permits an insulin release in response to D-glyceraldehyde, (D-GA; a glycolytic fuel) and L-leucine or alpha-ketoisocaproic acid (alpha-KIC) (non-glycolytic fuels), which alone are not initiators of insulin release in this species. 2. The "permissive" effect of D-glucose was also observed in the presence of D-mannose (which, as shown herein, is not insulinotropic alone). 3. The specificity of glucose for this "permissive" effect was, therefore, subsequently questioned in the presence of 10 mM alpha-KIC by substituting various glycolytic and non-glycolytic fuels to glucose. 4. D-GA (at 5 and 15 mM), D-mannose (30 and 50 mM), or the association of L-glutamine + L-asparagine permitted an insulin release in response to alpha-KIC. 5. The response was, however, delayed with D-GA, only occasionally with 50 mM D-mannose, and required high concentrations and was delayed in the presence of L-glutamine + L-asparagine as compared to that obtained with 14 mM D-glucose + alpha-KIC. 6. In conclusion, the threshold of fuel-induced insulin release is much higher in the chicken than in mammals and this threshold is most efficiently lowered by glucose.


A relationship between circulating natural glucocorticoids and the mechanical responses of the heart in atricial and precocial rodents.

1. A comparison was made of the mechanical performance of heart muscle from mouse, an atricial mammal, with corticosterone as glucocorticoid and spiny mouse (Acomys cahirinus), a precocial mammal, with cortisol as glucocorticoid. 2. Force-frequency responses were negative in mouse and positive in spiny mouse. 3. During recovery, there was a gradual increase and an overshoot in the mouse, while in the spiny mouse there was an initial enhanced response, diminishing gradually with time. 4. High calcium concentration inhibited contractile tension in mouse heart, while it was positively inotropic in spiny mouse heart. Changes in the concentration of calcium did not change the patterns of force-frequency response. 5. Lowering the experimental temperature increased the time course and amplitude of the tension curve. However, various parameters exhibited different temperature sensitivity. 6. There was a significant difference in the levels of circulating cortisol between male and female spiny mice. 7. It is proposed that the differences in the mechanical responses of mouse and spiny mouse hearts may be explained in terms of the effects of the specific glucocorticoid hormone on the development of the sodium-calcium exchanger.


The changes in body temperature, oxygen consumption, CO2 production and muscle protein turnover rate by selection for body size in Japanese quail, Coturnix coturnix japonica.

1. Body temperature, oxygen consumption, CO2 production and muscle protein degradation rate were measured in the three quail lines selected for body size, a random bred line (RR) and two lines selected for large (LL) or small (SS) body size. 2. The body temperature at 15 weeks of age was highest for small body size line and lowest for large body size line. 3. The body temperature, oxygen consumption and CO2 production of females were significantly higher than that of males. 4. The fractional degradation rate of muscle protein of SS, RR and LL lines were measured as 2.4, 1.6 and 1.2% per day in male, and 2.6, 1.7 and 1.4% per day in female.


Fibre digestion and digesta retention time in guinea-pigs (Cavia porcellus), degus (Octodon degus) and leaf-eared mice (Phyllotis darwini).

1. Digestibilities of feed and turnover time (1/k), Transit time (TT) and mean retention time (MRT: 1/k + TT) of fluid and particle markers were measured in the guinea-pig (Cavia porcellus), degu (Octodon degus) and leaf-eared mouse (Phyllotis darwini) fed a diet containing 50% alfalfa. 2. The digestibility of fibre and the retention time of digesta were highest in the guinea-pig followed by the degu and lowest in the leaf-eared mouse. 3. The difference in the retention time of digesta, resulting from the variation in the digestibility of fibre, between the three animals can be considered to be related to their body mass.


Dietary supplementation of vitamin E fails to prevent the development of hyperoxic lung injury in the premature guinea pig.

1. The benefit of dietary vitamin E supplementation in preventing oxidative-induced lung injury was investigated. Three day preterm guinea pig pups were exposed to hyperoxic (85% O2) or normoxic (21% O2) conditions. The animals were fed either a standard low birthweight human infant formula milk (6.4 mg/l vitamin E), or a vitamin E supplemented milk (100 mg/l) for up to 7 days. 2. After 3 days vitamin E supplementation, plasma but not erythrocyte vitamin E concentrations were elevated, while following 7 days both plasma and erythrocyte vitamin E concentrations were significantly increased. 3. Lung and liver vitamin E concentrations were elevated at both 3 and 7 days. At 3 days the increase in lung vitamin E was oxygen-dependent, suggesting that the lung increases uptake of vitamin E in response to oxidative stress. 4. Despite an increase in the vitamin E concentration of the lungs of preterm guinea pigs, no amelioration of the lung injury was observed. These results suggest that although vitamin E is a potent antioxidant, it is unable to protect adequately the lungs from reactive oxygen species in the absence of sufficient primary enzymatic antioxidant defences.


Effect of diet and essential amino acids gavage on young rat amino acid metabolism enzymes.

1. The effects of a high-fat, high-energy diet and essential plus semi-essential amino acid gavage on pup rats have been studied (60-65 animals). 2. The activities of alanine transaminase, adenylate deaminase, glutamine synthetase and serine dehydratase have been tested in liver and muscle. 3. Plasma was used for the estimation of proteins, urea, amino acids, glucose, lactate, 3-hydroxybutyrate and acetoacetate. 4. Liver and muscle glutamine synthetase activities are increased by diet and gavage administered. Hepatic serine dehydratase is inhibited by a cafeteria diet but activated by amino acid gavage. Adenylate deaminase is inhibited by diet and gavage in the liver, but gavage does not affect this enzyme activity in muscle. Liver alanine transaminase is increased by the diet; in the muscle, cafeteria diet and amino acid gavage showed the highest values for this enzyme. 5. In the plasma, the increase in lactate produced by the diet is inhibited by the amino acids provided. Cafeteria-fed pups showed lower urea levels and higher 3-hydroxybutyrate concentrations in the plasma. 6. Intracellular glucose is diminished by cafeteria diet. In contrast, the blood cell amino acid concentration increases with diet and gavage supplied.


Differential post-receptor responses of adenylate cyclase in white and brown adipose tissue membranes of rats fed high-energy diets.

1. Adenylate cyclase activity was determined in membranes of white and brown adipose tissue (WAT and BAT, respectively) from rats fed a high-energy diet (EXP group) vs those fed a nutritionally balanced one (CON group). 2. The isoproterenol- and guanine nucleotide-induced adenylate cyclase activity in WAT membranes of EXP rats was lower than that in CON rats. 3. Relative adenylate cyclase activity in like treated BAT membranes was higher in EXP than in CON rats. 4. It is concluded that feeding high-energy diets to rats induces similar post-receptor modifications of adenylate cyclase as found in genetic obese rodents.


A microcomputer program for determining turnover rates during non-steady state conditions: application to monoamine turnover.

A program to calculate the turnover rate of monoaminergic neurotransmitters was written for use on microcomputers. The program is based on non-steady state models, i.e., measuring the rate of change of levels of neurotransmitters or their metabolites after pharmacological inhibition of synthetic or degradative enzymes. Methods have been developed to estimate monoamine turnover by measuring the rate of decrease of catecholamine levels after treatment with alpha-methyl-p-tyrosine (AMPT), a tyrosine hydroxylase inhibitor, or the increase in serotonin levels after pargyline, an inhibitor of monoamine oxidase. This program is suitable for examining either linear increases or exponential decreases in monoamine levels. The program has been tested in our laboratory for the determination of serotonin, dopamine or norepinephrine turnover after hormonal or pharmacological manipulations. The program is readily adaptable for calculation of the turnover rate of other biological molecules under non-steady state conditions.


Intragastric nicotine protection against 40% ethanol injury in rat stomach. Role of ganglionic stimulation or blockade.

Intragastric nicotine (4 mg/kg) protects against 40% ethanol-induced gastric mucosal injury and raises mean blood pressure. We postulated that this protective effect was mediated by the ganglionic stimulatory property of nicotine and therefore could be abolished by ganglionic blockers. Rats were pretreated with intraperitoneal hexamethonium (10 mg/kg) or mecamylamine (2 mg/kg) to block peripheral or central autonomic ganglia, respectively. Intragastric vehicle or nicotine (4 mg/kg) was then administered. The total lengths of the linear gastric corpus mucosal lesions induced by intragastric 40% ethanol were measured by an unbiased observer using a caliper. The results showed that both intraperitoneal hexamethonium and mecamylamine pretreatments protected against 40% ethanol-induced gastric mucosal injury. Neither modified the protective effect of intragastric nicotine. The protective effect of hexamethonium and mecamylamine was associated with a significant increase in the volume of gastric mucus and gastric juice. The increase in the volume of gastric content (mucus and juice) was partially responsible for the protective effect of these ganglionic blockers. In a separate experiment, intraperitoneal nicotine (4 mg/kg) also protected against 40% ethanol-induced gastric mucosal injury and raised mean blood pressure. These data indicate that the protection against 40% ethanol-induced gastric mucosal injury is not unique to intragastric nicotine. Such protection can be induced by ganglionic blocking doses of hexamethonium and mecamylamine, or a ganglionic stimulatory dose of intraperitoneally administered nicotine. Whether ganglionic stimulation or blockade plays a role in the mechanism of intragastric nicotine protection, however, remains to be determined.(ABSTRACT TRUNCATED AT 250 WORDS)


Sequential histologic evaluations in collagenous colitis. Correlations with disease behavior and sampling strategy.

To evaluate the histologic manifestations of collagenous colitis and correlate histologic changes with disease behavior, 14 patients who had undergone sequential evaluations during 33 +/- 6 months of follow-up were studied. Two hundred twelve tissue specimens from all anatomic regions of the colon (mean, 15 +/- 3 samples per patient) were interpretated independently under code by two pathologists. Eight patients (57%) had histologic resolution after 14 +/- 4 months of empiric therapy and in only one of these (12%) did symptoms persist. Four patients (29%) had sequential histologic examinations from the same anatomic region that varied from classical collagenous colitis to inflamed mucosa without a thickened collagen band to normal mucosa. Eight patients (57%) had varying histologic findings from different anatomic regions during the same examination that ranged from classical collagenous colitis to increased inflammation with resolution of the collagen band to normal mucosa. Normal mucosa was found mainly in specimens from the rectosigmoid, and proctosigmoidoscopic examinations alone would have missed the diagnosis of collagenous colitis in 40% of cases. Pathologic interpretations were concordant in 171 of 212 instances (81%). We conclude that histologic resolution of collagenous colitis can occur and it is associated with loss of symptoms. The histologic features of collagenous colitis are distinctive, but they may be patchy and inconsistently sampled. Rectosigmoid biopsies underestimate the diagnosis.


Interrelationship between retinal ischaemic damage and turnover and metabolism of putative amino acid neurotransmitters, glutamate and GABA.

Conditions causing a reduction of oxygen availability (anoxia), such as stroke or diabetes, result in drastic changes in ion movements, levels of neurotransmitters and metabolites and subsequent neural death. Currently, there is no clinically available treatment for anoxia induced neural cell death resulting in drastic and permanent central nervous system dysfunction. However, there have been some exciting developments in experimentally induced anoxic conditions where several classes of drugs appear to significantly reduce neural cell death. This report aims to provide the foundations for understanding both the basic mechanisms involved in retinal ischaemic damage and experimental treatments used to prevent such damage. We discuss the normal release, actions and uptake of the fast retinal neurotransmitters, glutamate and GABA, in the vertebrate retina. Immunocytochemistry is used to demonstrate that both glutamate and GABA are found in the macaque retina. Following this is a discussion on how ischaemia may enhance neurotransmitter release or disrupt its uptake, thus causing an increase in extracellular concentration of these neurotransmitters and subsequent neuronal damage. The mechanisms involved in glutamate neurotoxicity are reviewed, because excess glutamate is the likely cause of retinal ischaemic damage. Finally, the mechanisms behind four possible modes of treatment of neurotransmitter toxicity and their advantages and disadvantages are discussed. Hopefully, further research in this area will lead to the development of a rational therapy for retinal, as well as cerebral ischaemia.


Perlman and Wiedemann-Beckwith syndromes: two distinct conditions associated with Wilms' tumour.

Though children with Perlman and Wiedemann-Beckwith syndromes have a number of features in common, the two conditions are probably separate entities. The distinction may not always be easy, however, partly because of the extreme rarity of Perlman syndrome, only nine cases of which have been reported so far. We report two siblings, initially diagnosed as having Wiedemann-Beckwith syndrome, in whom the correct diagnosis of Perlman syndrome was made only after an autopsy on the second child. By comparing and contrasting the features of Perlman and Wiedemann-Beckwith syndromes in this report we hope to make it easier to distinguish the two conditions.


Guinea pig hepatocyte alpha 1A-adrenoceptors: characterization, signal transduction and regulation.

Activation of guinea pig hepatocyte alpha 1-adrenoceptors increases phosphatidylinositol (PI) labeling, [3H]inositol phosphate production and phosphorylase activity. These adrenergic actions were not altered by pretreatment with chlorethylclonidine but were blocked by 5-methyl urapidil and prazosin (the former being 3- to 10-fold more potent than the latter), indicating that alpha 1A-adrenoceptors were involved. When the cells were incubated in buffer without calcium and containing EGTA, the alpha 1A-adrenergic stimulation of PI labeling was diminished but not abolished and that of phosphorylase was not affected. The alpha 1A-adrenergic effects were insensitive to pertussis toxin treatment. Phorbol myristate acetate inhibited the alpha 1A-adrenergic actions, although at relatively large concentrations, and also those of other agents such as angiotensin II and NaF. Our data clearly indicate that guinea pig hepatocytes express alpha 1A-adrenoceptors whose activation stimulates phosphoinositide turnover, via a pertussis toxin-insensitive process; the alpha 1A-adrenergic effects were at least partially independent of extracellular calcium.


Idazoxan down-regulates beta-adrenoceptors on C6 glioma cells in vitro.

Incubation of the C6 cells with 10 microM idazoxan (an alpha 2-adrenoceptor antagonist and putative antidepressant) for 5 days in vitro resulted in a 23% reduction of beta-adrenoceptor number and a 37% decrease in isoproterenol-induced cyclic AMP accumulation. In contrast, post-receptor stimulated cyclic AMP accumulation (by the use of forskolin or cholera toxin) was unaffected. The desensitization of the beta-adrenoceptor was accompanied by an increase in the KL/KH ratio for this receptor. Chronic in vitro treatment of C6 glioma cells with idazoxan did not significantly affect cholera or pertussis toxin catalyzed ribosylation of Gs and Gi/Go in these cells. Similarly, idazoxan did not alter either the basal levels of protein kinase C (PKC) alpha, or its cytoplasm to membrane translocation. These results suggest that idazoxan may have direct postsynaptic effects, the site of which may be at the level of receptor/G protein interaction.


L-2-amino-4-phosphonobutyrate (L-AP4) is an agonist at the type IV metabotropic glutamate receptor which is negatively coupled to adenylate cyclase.

Glutamate and L-AP4 inhibited forskolin-stimulated cyclic AMP (cAMP) production in baby hamster kidney (BHK) cells transfected with the type IV metabotropic receptor (mGluR4). In situ hybridization revealed a high level of mRNA for the mGluR4 in the entorhinal cortex, but not in the dentate gyrus. These data demonstrate that mGluR4 receptors are negatively coupled to the cAMP cascade, and suggest that the mGluR4 receptor may be the previously described presynaptic L-AP4 receptor.


Effects of drugs acting on Cl(-)-HCO3- and Na(+)-H+ exchangers on acid secretion in the rat gastric mucosa sheet preparation.

The effects of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), an inhibitor of the Cl(-)-HCO3- exchanger, and amiloride, an inhibitor of the Na(+)-H+ exchanger, on gastric acid secretion under basal conditions and after stimulation with bethanechol or dibutyryl cyclic AMP were studied in rat gastric mucosa sheet preparation. DIDS inhibited bethanechol-induced acid secretion in a dose-dependent manner, but amiloride had no effect. The stimulation of acid secretion by dibutyryl cyclic AMP plus 3-isobutyl-1-methylxanthine was also inhibited by DIDs, but not by amiloride. DIDS did not reduce basal acid secretion, and neither did amiloride. These results suggest that the Cl(-)-HCO3-exchanger in the basolateral membrane of the parietal cell plays an important role in stimulated gastric acid secretion and that the Na(+)-H+ exchanger is less important. In addition, these data show that DIDS inhibits stimulated gastric acid secretion irrespective of the secretagogue, but not basal gastric acid secretion.


Differential behavioural and neurochemical effects of competitive and non-competitive NMDA receptor antagonists in rats.

The behavioural and biochemical effects of the non-competitive N-methyl-D-aspartate (NMDA) receptor antagonists, dizocilpine and memantine, and the competitive NMDA receptor antagonist, CGP 39551, were investigated in rats. Systemic injections of dizocilpine (0.33 mg/kg) increased locomotion and rearing in an open field, whereas memantine (20 mg/kg) increased only locomotor activity. CGP 39551 (10 and 20 mg/kg) did not change open field activity. Dopamine (DA) metabolism--as measured by the ratio of dihydroxyphenylacetic acid/dopamine (DOPAC/DA)--increased in response to dizocilpine in the prefrontal cortex and the nucleus accumbens. Memantine enhanced DOPAC/DA in the prefrontal cortex, the nucleus accumbens and to a lesser degree in the posterior striatum. In contrast to non-competitive NMDA receptor antagonists, CGP 39551 did not increase DA metabolism of subcortical structures and even decreased DOPAC/DA in the prefrontal cortex. These results indicate that competitive and non-competitive NMDA receptor antagonists affect spontaneous locomotion differentially in rats. The biochemical data imply that the stimulant actions non-competitive NMDA receptor antagonists are at least partially due to activation of ascending dopaminergic systems. Potential mechanisms involved in the differential effects of both types of NMDA receptor antagonists are discussed.


Calcium channel blocking agents and potassium-stimulated release of glutamate from cerebellar slices.

The effects of calcium channel blockers and tetrodotoxin on the potassium-stimulated release of endogenous glutamate from rat cerebellar slices was assessed. Verapamil (10 microM), omega-conotoxin (1 and 10 microM) and cobalt (2 mM) all significantly decreased release. Amiloride (100 microM) and tetrodotoxin (0.5 and 1 microM) had no effect. The results suggest a role for N-type voltage-operated calcium channels in the potassium-stimulated release of glutamate.


Glucagon, insulin and somatostatin secretion in response to sympathetic neural activation in streptozotocin-induced diabetic rats. A study with the isolated perfused rat pancreas in vitro.

Changes in glucagon, insulin and somatostatin secretion induced by electrical splanchnic nerve stimulation were examined in rats treated with streptozotocin as neonates and as adults. In order to study the direct neural effects we used the isolated perfused rat pancreas with intact left splanchnic nerve in vitro. In normal rats splanchnic nerve stimulation causes significant decreases in insulin (30-40%) and somatostatin (30-50%) secretion at both 16.7 mmol/l and 1 mmol/l glucose concentrations. In the neonatal streptozotocin-diabetic rat splanchnic nerve stimulation at 16.7 mmol/l glucose decreased insulin secretion (14%) further than in the control rats (30%), however, somatostatin secretion did not decrease to the same extent. Similar results were also observed at the low (1 mmol/l) glucose concentration. On the other hand, percent decreases of insulin and somatostatin secretion induced by splanchnic nerve stimulation in the streptozocin-diabetic rats were similar to the values observed in the normal control rats. The glucagon secretion in response to splanchnic nerve stimulation at 16.7 mmol/l glucose from pancreatic Alpha cells in both types of induced diabetes is exaggerated, and the degree of exaggeration seems to parallel the severity of the hyperglycaemia. However, the splanchnic nerve stimulation-induced glucagon secretion at 1 mmol/l glucose was impaired in the streptozotocin-diabetic rats, but not in the neonatal streptozotocin-diabetic rats. These data suggest that the sensitivity of diabetic Alpha and Delta cells to sympathetic neural activation are blunted, whereas the sensitivity of Beta cells is enhanced in the diabetic animal model.


Transfection of murine P19S18 embryonal carcinoma cells with the oncogene neu induces an epithelioid phenotype.

An embryonal carcinoma cell line, P19S18, was transfected with the rat oncogene neu to investigate the function of its protein product, p185*, in a multipotential cellular environment. Levels of message for p185* were determined by in situ hybridization analysis and two highly expressing clones, PnnA and PnnB, were isolated. As demonstrated by indirect immunofluorescence and immunoprecipitation, these neu-transfected cells synthesized a full length rat p185*. The transfectants do not resemble typical embryonal carcinoma cells either before or after differentiation is induced by retinoic acid treatment. They are much larger, flatter, "epithelioid" cells. These cells have lost the expression of stage specific embryonic antigen-1 (SSEA-1), but do synthesize and assemble the basement membrane components laminin and fibronectin. These results suggest that expression of the neu oncogene in a multipotential cell line may induce the synthesis of proteins indicative of an epithelioid phenotype due to the presence of p185*.


[Polymorphism of untranscribed spacer rDNA as a molecular genetic marker in population studies of cattle].

Variable polymorphic patterns were detected using EcoRI-SalI fragment of bovine rDNA, including 3'-end of 28S rRNA gene with the adjacent portion of the transcribed spacer, as a probe for hybridization. Some features of these polymorphic patterns are similar to DNA fingerprints detected with the M13 probe. Bovine rDNA spacer polymorphism was used as a molecular genetic marker for population analysis of individual specific patterns of 4 cattle breeds with the help of the Jeffreys' method. It was supposed that the probability of identical fingerprints appearance could be the characteristics of heterogeneity of cattle populations. The observed length of polymorphic gragments ranged from 2000 to 6000 bp. The mean number of fragments per individual for all breeds was 15.05. The probability of identical patterns appearance was very high: from 1.18 x 10(-5) in ajshir's breed to 1.43 x 10(-7) in "white and black"s' breed. So, high probability seems to be dependent on the high allelic frequency and the way of breeding.


[Genetic differentiation of the Mongolian population. Comparative analysis of the geographic distribution of biochemical markers of genes and restriction fragment length polymorphism of nuclear DNA in the Mongolian population].

The geographical distribution of the gene frequencies from loci: Hp, Tf, Gc, Pi, AcP1, GLO1, EsD, 6-PGD, PGM1 and RFLP's of the nuclear DNA of the loci HBG-2 (HindIII), HBB (AvaII), ApoB (XbaI), D7S8 (PstI), LDLR (HincII) and AT-3 was analysed in the Mongolian population. These data revealed the homogeneity of 18 local groups in Mongolia and extremely low genetic differences measured by GST. There was no differences in the average GST values between protein markers and nuclear DNA markers.


A possible mechanism for the increase in brain tyrosine levels induced by cyanide in mice.

Subcutaneous administration of cyanide significantly increased blood tyrosine levels of mice in a dose dependent manner. Tyrosine aminotransferase activity in liver of mice was significantly decreased in the presence of cyanide (8, 10, 20, 40, 50, 65, 80 and 100 microM), also in a concentration-dependent manner, with a positive correlation between the percentage increase of blood tyrosine levels and the percentage decrease of hepatic tyrosine aminotransferase activity. These results suggest that the increased tyrosine levels induced in blood by cyanide may be related to its inhibition of tyrosine aminotransferase activity in the liver. Cyanide decreased hepatic ATP content and increased blood ammonia levels and brain tyrosine in a dose-dependent manner. As it is known that hyperammonaemia increases the uptake of neutral amino acids such as tyrosine into the brain from blood, the mechanism by which tyrosine levels increase in the brain may be based on increases of both tyrosine and ammonia levels in blood.


[Ammonium bituminosulfonate (Ichthyol). Anti-inflammatory effect and inhibition of the 5-lipoxygenase enzyme].

Ammonium bituminosulphonate (Ichthyol) inhibits 5-lipoxygenase activity in human polymorphonuclear neutrophils. The inhibition is dose-dependent and occurs at non-cytotoxic concentrations of the drug. This results in a decreased release of Leukotriene B4 from polymorphonuclear neutrophils. Furthermore, when applied to the ear skin of AB/Bln mice pretreated with croton oil, Ichthyol reduces the inflammatory reaction.


[Intralesional treatment of classical Kaposi sarcoma with interferon-alpha].

A 71-year-old female patient with classic Kaposi sarcoma (sarcoma idiopathicum haemorrhagicum multiplex) of the lower legs and one hand was treated with intralesional recombinant interferon alpha 2b. After 6 consecutive weeks of intratumoral injection of 1-3 million IU interferon-alpha 2b (Intron A) in three marked lesions, complete remission of intralesionally treated and of untreated tumours was observed. Even 9 months after treatment, the patient is still in complete remission. We conclude that this low dose and well-tolerated treatment schedule with interferon-alpha 2b is effective and a good alternative to radiation or chemotherapy in Kaposi sarcoma.


Analysis and pharmacokinetics of cimaterol in growing Holstein steers.

Pharmacokinetic parameters for the beta 2-adrenergic agonist, cimaterol (CIM), were determined in growing Holstein steers. Compartmental analysis was used after measurement of CIM in body fluids by affinity chromatography and HPLC using UV detection. Recoveries from spiked plasma and urine standards were 70 +/- 1.2% and 68 +/- 1.1%, respectively. The minimum detection level in plasma was 1 ng/mL and the average CV was 5.1% for concentrations that ranged from 1 to 30 ng/mL. Four steers (276 +/- 24 kg) received 15 mg of CIM by bolus intravenous injection. Plasma CIM levels declined in a biphasic manner with half-lives of 2.5 min for the distribution phase and 54 min for the elimination phase. A two-compartment open model was used to describe the disappearance of CIM and the following pharmacokinetic parameters were obtained: central compartment volume (Vc) = .76 L/kg, apparent volume of distribution (Vd) = 4.1 L/kg, and transfer rate constants from the central to peripheral compartment (k12) = .177/min, from the peripheral to central compartment (k21) = .054/min and elimination from the central compartment (kel) = .074/min. After 8 h, total urinary CIM accounted for only 18.3% of the administered dose. Results suggest that circulating concentrations of CIM in growing steers are influenced by its accumulation in an unidentified peripheral pool and its conversion into unknown metabolite(s) before elimination.


Changes in activities of lipogenic enzymes in adipose tissue and liver of growing goats.

The lipogenic capacity of omental adipose tissue and liver was measured in vitro from samples obtained at slaughter from 33 young male goats. The animals were slaughtered either on the day of weaning (d 0) or 2, 14, or 56 d after weaning. Ages at weaning were 4 wk (early weaning) or 6 or 8 wk (late weaning). Blood samples from the jugular vein were taken before slaughter to measure the concentrations of plasma glucose and nonesterified fatty acids. There was a 30% decrease in glucose concentration after weaning. Nonesterified fatty acid concentration increased fourfold between d 0 and 2 after weaning. By d 14 after weaning, nonesterified fatty acids returned to basal concentration. The lipoprotein lipase (LPL) activity of adipose tissue declined markedly (90%) on d 2 after weaning. Lipoprotein lipase activity returned to preweaning values by d 56 after weaning in those goats that had ad libitum access to feed. In adipose tissue, nicotinamide adenine dinucleotide phosphate (NADP)-malate dehydrogenase activity fell by only 17% by d 2 after weaning and to 63% by d 14 after weaning. Lipoprotein lipase activity was closely related to metabolizable energy intake the day before slaughter. Acetyl-coenzyme A carboxylase activity was low in adipose tissue and it increased only slightly by d 56 after weaning. The data indicated that LPL played a preponderant role in the restoration of lipid stores after weaning. High NADP-malate dehydrogenase activity together with a high concentration of plasma glucose by d 56 after weaning suggested that this enzyme activity could be enhanced by high glucose availability in goat kids. Activities of lipase, acetyl-coenzyme A carboxylase, NADP-malate dehydrogenase, and glucose-6-phosphate dehydrogenase in liver were essentially unaffected by weaning. The extent and rapidity of change of lipogenic enzymes of goat kids was much more pronounced in adipose tissue than in liver.


Pulmonary vasodilation by inhaled nitric oxide after endothelial injury.

Inhaled nitric oxide gas (NO) has recently been shown to reverse experimentally induced pulmonary vasoconstriction. To examine the effect of free radical injury and methylene blue exposure on inhaled NO-induced pulmonary vasodilation we studied ventilated rabbit lungs perfused with Krebs solution containing 3% dextran and indomethacin. When NO gas (120 ppm) was added to the inhaled mixture for 3 min, the elevated pulmonary arterial perfusion pressure (Ppa) induced by the thromboxane analogue U-46619 was significantly reduced [8 +/- 2 (SE) mmHg]. Acetylcholine similarly reduced Ppa (9 +/- 1 mmHg). After free radical injury and methylene blue exposure, inhaled NO again produced significant vasodilation (5 +/- 1 and 9 +/- 2 mmHg, respectively), but acetylcholine resulted in an increase in Ppa (-9 +/- 3 and -4 +/- 1 mmHg, respectively). These data demonstrate that pulmonary vasodilation produced by inhaled NO is unaffected by free radical injury or methylene blue in the intact lung despite concomitant reversal of acetylcholine-induced vasodilation.


Effects of nebivolol stereoisomers on the action of adrenaline on blood pressure, heart rate and blood levels of noradrenaline and DOPEG.

1. Nebivolol (a new beta 1-adrenoceptor blocking drug) has particular effects on the cardiovascular system, i.e. it induces hypotension without affecting cardiac function or increasing peripheral vascular resistances. In this study, we aimed to evaluate the effects of nebivolol and its stereoisomers on the actions of adrenaline (AD) at the cardiovascular level as well as at the prejunctional level (as ascertained by modification of noradrenaline (NA) and dihydroxyphenylglycol (DOPEG) plasma levels in the anaesthetized dog. 2. AD infusion (0.1 micrograms kg-1 min-1) did not induce statistically significant changes in mean blood pressure and heart rate; it caused a pronounced and sustained rise of AD levels, no significant alterations in NA levels and a marked, progressive and sustained increase in DOPEG levels. 3. Mean blood pressure was not affected by any of the nebivolol isomers. d- and dl-nebivolol in the two doses used (0.3 and 0.03 mg kg-1 in 15 min) caused a significant and dose-dependent decrease in heart rate. Plasma levels of AD and NA were not changed by any of the nebivolol isomers tested. However, all of them significantly reduced the increase in plasma levels of DOPEG induced by adrenaline infusion. 4. We conclude (1) that AD infusion in the dog facilitates NA release and that DOPEG is a good index of this effect; and (2) nebivolol appears to act at the prejunctional level, reducing the increase in NA release induced by adrenaline, as shown by the effect on DOPEG plasma levels.


A one-tube, one manipulation RT-PCR reaction for detection of Ross River virus.

A sensitive, single tube reverse transcription-polymerase chain reaction (RT-PCR) protocol for the detection of Ross River virus (RRV) is described. All components necessary for both reverse transcription and PCR were combined in a single tube, and reverse transcription and PCR carried out sequentially in a single, non-interrupted thermal cycling program. The antisense oligonucleotide from the two primers selected for use in the PCR also served to prime specifically for the reverse transcription. The 549 bp product was detected by electrophoresis and ethidium bromide staining. The detection limit using this system was 18 fg of purified viral RNA or 1.3 pfu of whole virus. Greater sensitivity cannot reasonably be expected unless a more sensitive method than electrophoresis and ethidium bromide staining is used for PCR product detection, such as nested PCR or hybridisation with labelled probe. This PCR detection system will be adapted for detection of RRV in mosquito populations for virus surveillance programs.


Impact of chronic oral H2-antagonist therapy on left ventricular systolic function and exercise capacity.

It has been suggested that H2-antagonists may adversely affect left ventricular systolic function. To assess the effects of cimetidine, famotidine, and ranitidine on exercise capacity and left ventricular systolic function, the authors conducted a randomized, double-blind, four-way crossover study in 15 healthy male volunteers with placebo control. Each subject underwent a maximal upright treadmill exercise test, aerobic metabolic assessment, and two-dimensional stress echocardiography on six separate occasions. The initial two treadmill exercise tests with aerobic metabolic assessment and stress echocardiography were performed to minimize the learning effect. In the final four evaluations, subjects were randomized to receive 7 days of oral treatment with cimetidine 400 mg twice daily, famotidine 40 mg daily, ranitidine 150 mg twice daily, and placebo. A comparison of exercise tests, aerobic metabolic assessment, and stress echocardiography results found no significant differences between any of the H2-antagonists and placebo. In addition, there were no significant differences in test results between cimetidine, famotidine, and ranitidine. Specifically, exercise treadmill time, maximal oxygen consumption, respiratory quotient, maximal exercise systolic and diastolic blood pressure, maximal exercise heart rate, left ventricular end-diastolic dimension, left ventricular end-systolic dimension, and ejection fraction were not different between treatments. The authors conclude that 7 days of oral treatment with cimetidine, famotidine, or ranitidine has no deleterious effect on exercise capacity or left ventricular systolic function in healthy subjects.


Benzodiazepines and other psychotropic drugs abused by patients in a methadone maintenance program: familiarity and preference.

Physicians often face the problem that they have to treat anxiety and insomnia in patients who are dependent on narcotics or other substances. Reports about different reinforcing properties of different benzodiazepines and increasing concern about their misuse, often in combination with other drugs, call for studies that help to establish recommendations for choosing psychopharmacologic agents in the treatment of these patients. Opiate addicts enrolled in a methadone maintenance program were interviewed about their subjective "liking" of all psychotropic substances with which they had personal experience. The results confirm previous reports that certain benzodiazepines, especially flunitrazepam and diazepam, stand out from others in terms of positive reinforcing properties. Overall, the attractiveness of benzodiazepines as drugs of abuse for poly-drug abusers is lower than that of other sedative/hypnotics.


Encapsulation and pilus formation of Bacteroides spp. in normal flora abscesses and blood.

The presence of encapsulated and piliated Bacteroides spp. (mostly Bacteroides fragilis and melaninogenicus groups was investigated in isolates from blood, abscesses and normal flora. Of the strains of Bacteroides spp. isolated 45 of 54 (83%) recovered from blood and 31 of 40 (78%) found in abscesses were encapsulated. In contrast, only seven of 71 (10%) similar strains isolated from the faeces or pharynx of healthy persons were encapsulated (P < 0.001). Pili were observed in three of 54 (6%) of strains isolated from blood, 30 of 40 (75%) of those recovered from abscesses (P < 0.001), and 49 of 71 (69%) of those found in normal flora (P < 0.001). The predominance of encapsulated forms in all strains of B. fragilis and B. melaninogenicus in blood as well as in abscesses suggests an increased virulence of these compared with non-encapsulated isolates. In contrast, the presence of pili in Bacteroides spp. recovered mostly from abscesses and normal flora suggests that this structure may play a role in the ability of these organisms to adhere to mucous membranes and may interfere with their ability to spread systemically. These findings illustrate the morphological differences that may be observed in Bacteroides spp. from various anatomical sites.


Typing by DNA probe of mycobacterial species isolated from patients with AIDS.

Restriction fragment length polymorphism (RFLP) types of Mycobacterium avium intracellulare and Mycobacterium kansasii isolated from patients with AIDS were examined. We demonstrate that one RFLP type is much more common than others which confirms previous findings. Carriage of individual RFLP types is constant over long periods of time. In addition, we document a disseminated infection in a patient with M. avium intracellulare of three RFLP types.


Efficacy of Bacillus sphaericus against the malaria vector Anopheles gambiae and other mosquitoes in swamps and rice fields in Zaire.

The microbial control of Anopheles gambiae and other mosquitoes with a granular formulation of Bacillus sphaericus (Vectolex) was evaluated in rice fields and swamps, located around the suburban region of Kingabwa-village in Kinshasa, Zaíre. Ten treatment cycles with 15-day intervals were carried out with the same application rate, 10 kg/ha, during the dry season (May to September 1991). The treatments reduced larval populations of An. gambiae by 98% after 48 h, but repetitive applications were required every 15 days to maintain control. The persistence of B. sphaericus spores was more apparent in rice fields than in swamps. A significant reduction in nuisance biting by Culex quinquefasciatus and Mansonia uniformis was observed. For An. gambiae, a decrease of 13.6% in human biting was noted during the post-treatment period. The entomological inoculation rate was reduced from 0.238 to 0.143. The efficacy of B. sphaericus does not appear to offer outstanding potential for control of An. gambiae in rice fields and swamps and seems to be limited due to different factors tied to ecology and natural conditions in the fields.


A fabric body light trap for sampling mosquitoes.

A Fabric Body light trap has been developed that has unique features of a cloth body, folding rainshield and a modified folding net. These features allow the light trap to be reduced in size and weight for transportation and storage while retaining the same operational characteristics as the Solid State Army Miniature (SSAM) light trap.


Reduced mosquito production in cemetery vases with copper liners.

Water-holding stone vases were sampled in 4 central Florida cemeteries to compare the prevalence of mosquitoes in containers with and without metallic liners. Overall, immature mosquitoes were found in more than 60% of the vases lacking liners and in more than 50% of the vases with aluminum liners. Significantly fewer vases with copper liners were positive for mosquitoes. High mortality and a lack of development were observed in a field test involving the introduction of Aedes aegypti larvae into stone vases with copper liners.


A survey of larval mosquitoes on Kume Island, Ryukyu Archipelago, Japan.

Mosquito collections were carried out during May 1991 in Kume Island, Ryukyu Archipelago. Eighteen species of mosquitoes in 8 genera were collected; 11 species were new records for Kume Island. Characteristics of the larval habitats of these species are also described.


Effect of antibiotic treatment on mycelial growth of isolates of the mosquito pathogenic fungus Lagenidium giganteum (Oomycetes: Lagenidiales) from Australia, Colombia and United States.

Bacteria collected in mosquito breeding ponds were evaluated for resistance to streptomycin, chloramphenicol, penicillin and trimethoprim. Mycelial growth of Lagenidium giganteum isolates from Australia, United States and Colombia were evaluated in PYG media containing one antibiotic or a mixture of these compounds. Media containing chloramphenicol reduced mycelial growth of most of the isolates. The antibiotic mixtures and penicillin-streptomycin penicillin-trimethoprim did not significantly affect mycelial growth of the isolates; however, the later substantially reduced bacterial contamination.


A comparison of T4:T8 lymphocyte ratio in the periodontal lesion of healthy and HIV-positive patients.

Previous reports describe a characteristic, rapidly progressive, periodontitis that is unique to patients who are seropositive for HIV antibody (Western blot +). The purpose of this study was to compare the T4 and T8 lymphocyte subpopulations in the peripheral blood and periodontal lesions of these HIV patients with those of healthy controls. T-cell subsets in peripheral blood were quantified by flow cytometry. The values from this analysis were used to calculate the peripheral T4:T8 lymphocyte ratio for each patient. Gingival tissue (papilla) was obtained from 8 HIV+ patients and from 6 healthy HIV- control patients during routine gingival surgery. The T-cell subpopulations in the gingival tissue were determined using serial cryostat sections that were labeled with monoclonal antibodies for T4 and T8 cells and developed using an avidin-biotin-peroxidase system. Six sections were taken from each of the 14 tissue specimens (one per patient). The sections were examined at 450 x and the mean number of T4 and T8 cells calculated for each section. These mean values were then used to determine the T4:T8 lymphocyte ratio for each tissue specimen. The peripheral blood analysis revealed a mean serum T4:T8 ratio of (2.07 +/- 0.455) for the controls and (0.58 +/- 0.26) for the HIV patients. The significantly lower T4:T8 ratio in HIV patients is consistent with their diagnosis. Although the results indicated that the mean T4:T8 lymphocyte ratio in the gingiva of controls was highly variable (2.70 +/- 1.344), the gingiva of HIV patients consistently exhibited a complete absence of T-cells.(ABSTRACT TRUNCATED AT 250 WORDS)


Opioid receptor affinity for agonist-antagonist analgesics.

Analgesic medications are distributed to a variety of receptors within the central nervous system. Activity at these receptors (mu 1, mu, sigma, delta, kappa) results in both the beneficial pain-relieving effects of analgesics as well as undesirable side effects. The mixed agonist-antagonist class of analgesics offers the potential benefit of greater receptor site selectivity while diminishing the incidence of adverse sequelae, such as respiratory depression. Traditionally, it has been suggested that mixed agonist-antagonist medications may be associated with decreased analgesic effectiveness. However, newer agents of this mixed class may result in effective analgesia while diminishing the incidence of side effects.


Role of B13 Glu in insulin assembly. The hexamer structure of recombinant mutant (B13 Glu-->Gln) insulin.

The assembly of the insulin hexamer brings the six B13 glutamate side-chains at the centre into close proximity. Their mutual repulsion is unfavourable and zinc co-ordination to B10 histidine is necessary to stabilize the well known zinc-containing hexamers. Since B13 is always a carboxylic acid in all known sequences of hexamer forming insulins, it is likely to be important in the hormone's biology. The mutation of B13 Glu-->Gln leads to a stable zinc-free hexamer with somewhat reduced potency. The structures of the zinc-free B13 Gln hexamer and the 2Zn B13 insulin hexamer have been determined by X-ray analysis and refined with 2.5 A and 2.0 A diffraction data, respectively. Comparisons show that in 2Zn B13 Gln insulin, the hexamer structure (T6) is very like that of the native hormone. On the other hand, the zinc-free hexamer assumes a quaternary structure (T3/R3) seen in the native 4Zn insulin hexamer, and normally associated only with high chloride ion concentrations in the medium. The crystal structures show the B13 Gln side-chains only contact water in contrast to the B13 glutamate in 2Zn insulin. The solvation of the B13 Gln may be associated with this residue favouring helix at B1 to B8. The low potency of the B13 Gln insulin also suggests the residue influences the hormone's conformation.


Morphologic changes in cultured astrocytes after exposure to glutamate.

Cultured astrocytes, exposed to glutamate at a dose that is generally neurotoxic in vitro (1 mM), exhibit transient swelling in the absence of cell death. In the present study, we further characterize this response by examining the distribution of intermediate filaments and evaluating cellular ultrastructure in primary cultures of astrocytes after exposure to 1 mM glutamate. In addition, cellular swelling was determined using the nonmetabolizable hexose 3-O-methyl [14C]-glucose (3-MG). Glial fibrillary acidic protein (GFAP) was immunolocalized at the light microscopic level to study the distribution of intermediate filaments. GFAP was immunolocalized to a fine cytoskeletal network in control cultures. Four to 24 h after exposure to glutamate, this detailed localization was replaced by a diffuse, uneven pattern of immunoreactivity. The most prominent ultrastructural changes were identified at 4 and 8 h after glutamate exposure. Nucleoli underwent transformation from a normal compact appearance to a markedly dispersed state. The cell body typically exhibited cytoplasmic lucency, swollen mitochondria, and dilated cisterns. Intermediate filaments within cellular processes appeared widely spaced in comparison to the controls. These ultrastructural changes coincided with findings of increased intracellular water space as determined with 3-MG. These findings demonstrate that astrocytes exposed to 1 mM glutamate exhibit transient morphologic changes that not only suggest cellular swelling but also define a more diverse response that is reflected in the altered immunolocalization of GFAP and in unique changes in the nucleolus.


Selective effect of chronic lead ingestion on tyrosine hydroxylase activity in brain regions of rats.

Alterations of tyrosine hydroxylase activity in various regions of brain from rats postnatally exposed to lead were tested. Three groups of animals were prepared; (1) Rats exposed to lead at a low dose (0.05% lead acetate, PbAc); (2) Rats exposed to lead at a high dose (0.2% PbAc); (3) Age-matched normal control rats. At 2, 4, 6, and 8 weeks of age, weight of brain and body, and concentrations of lead in whole brain of animals in each group were measured. Activities of tyrosine hydroxylase and Na(+)-K+ ATPase were also measured at the same ages in 4 brain regions of each animal. Body weight gain was decreased after 6 weeks of age in rats exposed to lead at a high dose. Concentrations of lead in whole brain were increased from 0.37 to 0.83 (ng/mg wet tissue) in these animals. Exposure of rats to lead generally increased tyrosine hydroxylase activity and decreased Na(+)-K+ ATPase activity. However, changes of tyrosine hydroxylase activity were detected without concomitant changes of Na(+)-K+ ATPase activity in pons-medulla at 2 weeks of age and telencephalon at 6 weeks of age in rats exposed to lead at a low dose, and in midbrain at 4 and 6 weeks of age in rats exposed to lead at a high dose. These data imply that catecholaminergic nervous system in the brain regions described above could be selectively affected by lead.


Effects of hexamethonium on bradycardiac responses to brain ischemia in the rabbit.

The present study investigated the bradycardiac responses to brain ischemia for approximately 30s before and after intravenous administration of hexamethonium (C6, 15 mg/kg) in urethane-anesthetized spontaneously breathing rabbits. The brain ischemia was performed by clamping both common carotid arteries in rabbits whose vertebral arteries were previously occluded. The brain ischemia caused bradycardia, pressor response and apnea. Administration of C6 blocked the bradycardia evoked by brain ischemia and reduced pressor response at the initial period after the onset of brain ischemia. The brain ischemia-induced apnea was not significantly altered by C6-treatment. In a separate series of experiments, we examined the effects of C6 on the response of heart rate (HR) to vagal stimulation in rabbits following unilateral vagotomy. Electrical stimulation of the peripheral end of the cut left vagus nerve that selectively activated myelinated fibers caused the bradycardia and this effect was entirely blocked by administration of C6. When the intensity of stimulus to activate both myelinated and non-myelinated fibers was increased, part of the bradycardia was retained following C6 administration. These results suggest that the brain ischemia-induced bradycardia is totally mediated through the activation of myelinated efferent fibers in the vagus nerve.


[int-2 and c-erbB-2 gene amplification in urological cancers].

We analyzed the alteration of int-2, c-erbB-2 and EGFR genes in 32 cases of transitional cell carcinoma of the urinary tract, 15 cases of renal cell carcinoma and 14 cases of prostatic carcinoma by Southern blot hybridization method. Three- to 12 fold amplification of int-2 gene was observed in 4 (12.5%) of 32 transitional cell carcinomas. Of these 4 cases 3 were G3 tumor with muscle invasion and the remaining was G1, pTa tumor with subsequent recurrence of multiple tumors. The other 2 cases (6.3%) with invasive transitional cell carcinoma showed amplification of c-erbB-2 gene. Neither amplification nor gross rearrangement of EGFR gene was detected in transitional cell carcinoma. On the other hand, renal cell carcinomas and prostatic carcinomas had neither amplification nor gross rearrangement of these 3 genes. These results suggest that the int-2 gene located in chromosome locus 11q13 and the c-erbB-2 gene have a specific role in carcinogenesis and in progression of transitional cell carcinoma through their gene amplifications.


NMDA-, but not kainate- or quisqualate-dependent increases in cerebellar cGMP are dependent upon monoaminergic innervation.

As previously reported, intracerebellar injections of D-serine, quisqualate and kainate elevated mouse cerebellar cGMP levels. Similarly, activation of endogenous excitatory amino acid utilizing neurons with harmaline or pentylenetetrazole also increased cerebellar cGMP. We previously have demonstrated that the harmaline- and pentylenetetrazol-dependent cGMP increases are NMDA receptor mediated. In this report we further demonstrate that NMDA-dependent increases in cGMP are dependent upon monoaminergic innervation of the cerebellum, while kainate- and quisqualate-dependent cGMP increases are independent of such innervation.


A specific enzyme assay for aminopeptidase M in rat brain.

A specific enzyme assay for aminopeptidase M (APM) activity on rat brain membranes has been developed through selective use of enzyme inhibitors. Amastatin was the most potent inhibitor (amastatin > actinonin > MDL73347 > bestatin) for purified porcine kidney APM, giving 98% inhibition at a 6 microM concentration, while actinonin, yielded only 57% inhibition at this concentration. Puromycin (10 microM) was used to inhibit puromycin-sensitive aminopeptidase activity in the rat brain membrane preparation. Puromycin (10 microM) had only a slight effect on the Km of porcine kidney APM, and had negligible effect on APM velocity at the high substrate concentration (2 mM) used in the APM assay. The assay produced a linear accumulation of product for increasing amount of rat brain membranes used, and for increasing incubation time. The Km of APM on rat brain membranes for L-Leucine-p-nitroanilide (0.383 mM) was similar to the Km of purified porcine kidney APM (0.558 mM). APM-activity, involved in the metabolism of several biologically important neuropeptides in different brain regions, can be specifically measured with this enzyme assay.


Studies on novel pili from Shigella flexneri. I. Detection of pili and hemagglutination activity.

Pili were detected using electron microscopy in clinical isolates of Shigella flexneri which had been continuously subcultivated in liquid media. Morphologically, the pili appeared as thin, flexible, cylindrical structures of up to 2-5 microns in length and about 3-5 nm in diameter. Two strains showed mannose-resistant (MR) hemagglutination to fresh fowl erythrocytes (type 4), and one to tannic acid-treated horse erythrocyte (type 3) pili. These pili are novel and different from the mannose-sensitive (MS) type 1 pili described by Duguid and Gillies.