Covert attention in touch: behavioral and ERP evidence for costs and benefits

To investigate the mechanism underlying tactile spatial attention, reaction times (RTs) and event-related potentials (ERPs) were recorded in response to mechanical stimuli delivered to the hands. At the start of each trial cues indicated either the correct (valid) or incorrect (invalid) tactile stimulus location or were uninformative (neutral). RT costs (suppression of invalid compared to neutral trials) were found to be larger than benefits (enhancement of valid compared to neutral trials). ERPs showed that costs and benefits contribute equally to attentional modulations of the somatosensory N140 component, whereas these were largely due to costs at longer latencies. These results differ from the pattern of attentional ERP modulations previously found for vision and audition, where costs precede benefits, and therefore suggest that the mechanisms of attentional selectivity in touch might be different from attentional processes in other modalities.     

Antimicrobial susceptibility testing of aquatic bacteria: quality control disk diffusion ranges for Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658 at 22 and 28 degrees C

Quality control (QC) ranges for disk diffusion susceptibility testing of aquatic bacterial isolates were proposed as a result of a multilaboratory study conducted according to procedures established by the National Committee for Clinical Laboratory Standards (NCCLS). Ranges were proposed for Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658 at 22 and 28 degrees C for nine different antimicrobial agents (ampicillin, enrofloxacin, erythromycin, florfenicol, gentamicin, oxolinic acid, oxytetracycline, ormetoprim-sulfadimethoxine, and trimethoprim-sulfamethoxazole). All tests were conducted on standard Mueller-Hinton agar. With >/=95% of all data points fitting within the proposed QC ranges, the results from this study comply with NCCLS guidelines and have been accepted by the NCCLS Subcommittee for Veterinary Antimicrobial Susceptibility Testing. These QC guidelines will permit greater accuracy in interpreting results and, for the first time, the ability to reliably compare susceptibility test data between aquatic animal disease diagnostic laboratories.     

Deviant expression of Rab5 on phagosomes containing the intracellular pathogens Mycobacterium tuberculosis and Legionella pneumophila is associated with altered phagosomal fate

The intracellular human pathogens Legionella pneumophila and Mycobacterium tuberculosis reside in altered phagosomes that do not fuse with lysosomes and are only mildly acidified. The L. pneumophila phagosome exists completely outside the endolysosomal pathway, and the M. tuberculosis phagosome displays a maturational arrest at an early endosomal stage along this pathway. Rab5 plays a critical role in regulating membrane trafficking involving endosomes and phagosomes. To determine whether an alteration in the function or delivery of Rab5 could play a role in the aberrant development of L. pneumophila and M. tuberculosis phagosomes, we have examined the distribution of the small GTPase, Rab5c, in infected HeLa cells overexpressing Rab5c. Both pathogens formed phagosomes in HeLa cells with molecular characteristics similar to their phagosomes in human macrophages and multiplied in these host cells. Phagosomes containing virulent wild-type L. pneumophila never acquired immunogold staining for Rab5c, whereas phagosomes containing an avirulent mutant L. pneumophila (which ultimately fused with lysosomes) transiently acquired staining for Rab5c after phagocytosis. In contrast, M. tuberculosis phagosomes exhibited abundant staining for Rab5c throughout its life cycle. To verify that the overexpressed, recombinant Rab5c observed on the bacterial phagosomes was biologically active, we examined the phagosomes in HeLa cells expressing Rab5c Q79L, a fusion-promoting mutant. Such HeLa cells formed giant vacuoles, and after incubation with various particles, the giant vacuoles acquired large numbers of latex beads, M. tuberculosis, and avirulent L. pneumophila but not wild-type L. pneumophila, which consistently remained in tight phagosomes that did not fuse with the giant vacuoles. These results indicate that whereas Rab5 is absent from wild-type L. pneumophila phagosomes, functional Rab5 persists on M. tuberculosis phagosomes. The absence of Rab5 on the L. pneumophila phagosome may underlie its lack of interaction with endocytic compartments. The persistence of functional Rab5 on the M. tuberculosis phagosomes may enable the phagosome to retard its own maturation at an early endosomal stage.     

Effects of gastrointestinal candidiasis, antibiotics, dietary arabinitol, and cortisone acetate on levels of the Candida metabolite D-arabinitol in rat serum and urine.

We studied the effects of gastrointestinal (GI) colonization by Candida albicans, dietary arabinitol, intragastric antibiotics, and cortisone on levels of the Candida metabolite D-arabinitol in rat serum and urine. Rats given conventional laboratory chow, intragastric gentamicin and chloramphenicol, and 6.0 x 10(8) live C. albicans B311 blastoconidia by gavage had minimal invasive GI disease and no more DL-arabinitol in the urine than controls given killed C. albicans. However, colonized and uncolonized rats given intragastric antibiotics had transiently higher urine arabinitol levels than the corresponding controls given saline. Rats given conventional laboratory chow (which contained 50 micrograms of arabinitol per g) had higher serum and urine arabinitol levels than rats given no dietary arabinitol, but the differences were less than expected. Moreover, intragastric antibiotics did not cause increased arabinitol excretion in rats given no dietary arabinitol. Rats given intragastric antibiotics and live C. albicans but no dietary arabinitol had no more arabinitol in their serum or urine than controls given antibiotics and killed C. albicans or saline and live or killed C. albicans. Lastly, cortisone acetate (10 mg/kg of body weight per day intramuscularly for 10 days) did not cause increased serum or urine arabinitol levels. We conclude that neither GI colonization by C. albicans nor cortisone should interfere with the usefulness of arabinitol as a marker for invasive candidiasis; antibiotics appear to increase arabinitol excretion by suppressing GI bacteria capable of consuming dietary arabinitol.     

Identified serotonergic neurons in the Tritonia swim CPG activate both ionotropic and metabotropic receptors

Although G-protein-coupled (metabotropic) receptors are known to modulate the production of motor patterns, evidence from the escape swim central pattern generator (CPG) of the nudibranch mollusk, Tritonia diomedea, suggests that they might also participate in the generation of the motor pattern itself. The dorsal swim interneurons (DSIs), identified serotonergic neurons intrinsic to the Tritonia swim CPG, evoke dual component synaptic potentials onto other CPG neurons and premotor interneurons. Both the fast and slow components were previously shown to be due to serotonin (5-HT) acting at distinct postsynaptic receptors. We find that blocking or facilitating metabotropic receptors in a postsynaptic premotor interneuron differentially affects the fast and slow synaptic responses to DSI stimulation. Blocking G-protein activation by iontophoretically injecting the GDP-analogue guanosine 5'-O-(2-thiodiphosphate) (GDP-beta-S) did not significantly affect the DSI-evoked fast excitatory postsynaptic potential (EPSP) but decreased the amplitude of the slow component more than 50%. Injection of the GTP analogues guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-S) and 5'-guanylyl-imidodiphosphate, to prolong G-protein activation, had mixed effects on the fast component but increased the amplitude and duration of the slow component of the DSI-evoked response and, with repeated DSI stimulation, led to a persistent depolarization. These results indicate that the fast component of the biphasic synaptic potential evoked by a serotonergic CPG neuron onto premotor interneurons is mediated by ionotropic receptors (5-HT-gated ion channels), whereas the slow component is mediated by G-protein-coupled receptors. A similar synaptic activation of metabotropic receptors might also be found within the CPG itself, where it could exert a direct influence onto motor pattern generation.     

Laser capture microdissection and real-time PCR for analysis of glomerular endothelin-1 gene expression in mesangiolysis of rat anti-Thy 1.1 and murine Habu Snake Venom glomerulonephritis.

Molecular analysis of pathologic changes in glomeruli requires methods allowing rapid and exact detection of alterations in gene expression. Here, we analyzed endothelin-1 (ET-1) mRNA expression in mesangiolytic glomeruli during the course of a rat and murine model of mesangioproliferative glomerulonephritis (GN). A novel method combining laser capture microdissection (LCM), which permits the precise removal of selected mesangiolytic glomeruli, with a highly sensitive real-time RT-PCR technique was used. Anti-Thy 1.1. GN was introduced in male Sprague-Dawley rats (1.0 mg/kg body weight of OX-7 IV) and Habu Snake Venom GN was introduced in C57BL6 mice (habu snake venom toxin 6 mg/kg body weight IV). The degree of mesangiolysis during both GNs was analyzed using a semiquantitative scoring system. Mesangiolytic glomeruli were microdissected at different days of the diseases (day 2, 6, and 12 in anti-Thy 1.1 GN and days 1, 3, 7, and 14 in Habu Snake Venom GN) and from normal control animals. After RNA extraction and cDNA synthesis, ET-1 gene expression was measured by real-time RT-PCR. In parallel, in anti-Thy 1.1. GN ET-1 mRNA expression was analyzed using semiquantitative nonradioactive in situ hybridization; ET-1 protein expression was investigated by immunohistochemistry. Mesangiolysis peaked at day 6 in anti-Thy1.1 GN and at day 1 in Habu Snake Venom GN. Mesangiolytic glomeruli were easily microdissected on cryostat sections in both models; quantification of mRNA with RT-PCR was reliable and reproducible. Glomerular ET-1 mRNA expression increased during the course of anti-Thy 1.1 GN and Habu Snake Venom GN peaked when mesangiolysis was most pronounced. This was seen by RT-PCR after glomerular LCM and by in situ hybridization; in parallel, glomerular ET-1 protein expression was increased. Combination of LCM and RT-PCR is a reliable method for quantification of localized gene expression in isolated renal structures. The above data argue for an important role of ET-1 in pathogenesis and/or repair of mesangiolysis in experimental mesangioproliferative GN.     

Synthetic amyloid beta-protein fails to produce specific neurotoxicity in monkey cerebral cortex.

Because progressive amyloid beta-protein (A beta P) deposition and surrounding neuritic dystrophy occur spontaneously in primates, we evaluated the in vivo effects of synthetic A beta P in monkey cortex. Experimental and control A beta P were stereotactically injected into multiple neocortical sites of adult rhesus monkeys in a vehicle of either artificial cerebrospinal fluid or acetonitrile. After 2 weeks or 3 months, injection sites were identified and characterized histologically and immunocytochemically. A beta P antibodies specifically detected the injected A beta P1-40 peptide. Serial sections stained with silver and antineurofilament protein demonstrated comparable degrees of degenerating neurons, dystrophic neurites, and axonal spheroids associated with both experimental and control peptide injections. Alz 50 staining was sparse or absent in all sites. We conclude that specific cellular changes closely resembling AD pathology were not detected in these experiments, and that control and experimental A beta P peptides produced indistinguishable effects. Methodological concerns regarding the in vivo modeling of A beta P bioactivity are discussed.     

Delayed responses to electrical stimuli reflect C-fiber responsiveness in human microneurography

The slowing of impulse conduction during the relative refractory period has often been used to assess activation of C-fibers, in particular, in human microneurography. This study aimed to evaluate the sensitivity of this method and the factors affecting it. Thirty cutaneous C-fibers were recorded from the peroneal nerves of healthy human subjects. Intracutaneous electrical stimulation in the receptive field at 4 s intervals, after some minutes of adaptation, induced spike discharges at constant latency. One or more conditioning stimulus pulses were interpolated at different intervals and the increase in latency after the subsequent regular pulse was assessed. The latency shift was found to depend on the number of interposed pulses, on the time interval between conditioning and conditioned stimulus, and on the conduction velocity of the C-unit. The increase in latency was larger with greater distance between stimulating and recording electrodes, indicating a contribution of the conductile membrane over its whole length. On the other hand, slowing was more pronounced, on average, in slower conducting C-units and conduction velocities were slower when recordings were performed more distally. These findings indicate that the slower terminal nerve branches contribute most to the latency increases. Even a single additional spike in between two regular pulses caused a reliable latency shift of 1.2±0.2 ms (mean ±SEM) and additional pulses lead to an approximately linear latency increase (2 pulses: 2.3±0.3 ms; 4 pulses: 5.9±0.7 ms). In contrast to the number of interposed stimuli, different intervals between interposed and regular stimuli had only a minor impact on the latency shifts. It is concluded that latency shifts are reliable indicators of C-fiber activation, being sensitive enough to detect even single spike responses. Furthermore, latency increases may be used as a relative measure of C-fiber activation, e.g., when comparing responses to stimuli of different strength.     

IgD production and other lymphocyte functions in HIV infection: immaturity and activation of B cells at different clinical stages.

T and B cell function, in particular IgD production in vitro, were studied across the spectrum of HIV infection in homosexual men and compared with seronegative homosexual and heterosexual male controls. Proliferation to phytohaemagglutinin (PHA) was reduced most strikingly in symptomatic HIV infection; it was also impaired in HIV seronegative homosexual men and there was no difference between these and asymptomatic HIV seropositives or those with persistent generalized lymphadenopathy (PGL). Spontaneous IgG and IgM production were increased in patients with PGL and Kaposi's sarcoma; pokeweed mitogen (PWM)-induced production of IgG and IgM was reduced in all HIV infected subjects. Spontaneous production of IgD was highest in asymptomatic HIV infection, with raised values also seen in PGL and AIDS with opportunist infection; IgD production was suppressed by PWM in the same groups. These data indicate an increase in circulating immature B cells. Markers of B cell immaturity and polyclonal activation are apparent to differing degrees at different stages of HIV infection.