PKCgamma contributes to a subset of the NMDA-dependent spinal circuits that underlie injury-induced persistent pain.

In previous studies we provided evidence that the gamma isoform of protein kinase C (PKCgamma) is an important contributor to the increased pain sensitivity that occurs after injury. Here we combined electrophysiological and behavioral approaches in wild-type and PKCgamma-null mice to compare the hyperexcitability of wide dynamic range neurons in lamina V of the spinal cord dorsal horn with the behavioral hyperexcitability produced by the same injury [application of a C-fiber irritant, mustard oil (MO), to the hindpaw]. Wild-type and null mice did not differ in their response to mechanical or thermal stimuli before tissue injury, and the magnitude of the response to the MO stimuli was comparable. In wild-type mice, MO produced a dramatic and progressive enhancement of the response of lamina V neurons to innocuous mechanical and thermal stimuli. The time course of the neuronal hyperexcitability paralleled the time course of the MO-induced behavioral allodynia (nocifensive behavior in response to a previously innocuous mechanical stimulus). Neuronal hyperexcitability was also manifest in the PKCgamma-null mice, but it lasted <30 min. By contrast, the behavioral allodynia produced by MO in the PKCgamma-null mice, although reduced to approximately half that of the wild-type mice, persisted long after the lamina V hyperexcitability had subsided. Because the MO-induced behavioral allodynia was completely blocked by an NMDA receptor antagonist, we conclude that PKCgamma mediates the transition from short- to long-term hyperexcitability of lamina V nociresponsive neurons but that the persistence of injury-induced pain must involve activity within multiple NMDA-dependent spinal cord circuits.

Performance characteristics of three serum iron and total iron-binding capacity methods in acute iron overdose.

Accurate serum iron and total iron binding capacity (TIBC) measurements may be useful in acute iron overdoses. Two alumina column TIBC methods were found to measure increased TIBC when free iron was present. A homogeneous TIBC method gave consistent results until iron concentrations exceeded 500 micrograms/dL (90 mumol/L), when it began to underestimate the TIBC. Serious iron overdoses require chelation therapy with deferoxamine. Iron recovery was reduced by up to 50% for all 3 methods with clinically achievable concentrations of deferoxamine 8,400 micrograms/dL (150 mumol/L). TIBC measurements by both alumina column methods were reduced by deferoxamine in the presence of free iron and unaffected when the iron concentration was less than the TIBC. The homogeneous TIBC method yielded falsely elevated results in the presence of free deferoxamine. Procedures that measure TIBC by addition of excess ferric iron followed by alumina adsorption are not suitable for monitoring TIBC in acute iron overdose. The homogeneous TIBC assay can be used in acute iron overdose but underestimates TIBC when iron concentrations exceed 500 micrograms/dL (90 mumol/L). None of the methods examined are useful for measuring iron or TIBC in the presence of deferoxamine.

Inflammation-induced up-regulation of protein kinase Cgamma immunoreactivity in rat spinal cord correlates with enhanced nociceptive processing.

Activation of various second messengers contributes to long-term changes in the excitability of dorsal horn neurons and to persistent pain conditions produced by injury. Here, we compared the time-course of decreased mechanical nociceptive thresholds and the density of protein kinase Cgamma immunoreactivity in the dorsal horn after injections of complete Freund's adjuvant in the plantar surface of the rat hindpaw. Complete Freund's adjuvant significantly increased paw diameter and mechanical sensitivity ipsilateral to the inflammation. The changes peaked one day post-injury, but endured for at least two weeks. In these rats, we recorded a 75-100% increase in protein kinase Cgamma immunoreactivity in the ipsilateral superficial dorsal horn of the L4 and L5 segments at all time-points. Electron microscopy revealed that the up-regulation was associated with a significant translocation of protein kinase Cgamma immunoreactivity to the plasma membrane. In double-label cytochemical studies, we found that about 20% of the protein kinase Cgamma-immunoreactive neurons, which are concentrated in inner lamina II, contain glutamate decarboxylase-67 messenger RNA, but none stain for parvalbumin or nitric oxide synthase. These results indicate that persistent changes in protein kinase Cgamma immunoreactivity parallel the time-course of mechanical allodynia and suggest that protein kinase Cgamma contributes to the maintenance of the allodynia produced by peripheral inflammation. The minimal expression of protein kinase Cgamma in presumed inhibitory neurons suggests that protein kinase Cgamma-mediated regulation of excitatory interneurons underlies the changes in spinal cord activity during persistent nociception.

Anatomical basis for cannabinoid-induced antinociception as revealed by intracerebral microinjections.

Cannabinoids suppress behavioral and neurophysiological responses to noxious stimuli in rodents when administered systemically. The purpose of this study was to extend previous studies of the site of cannabinoid analgesia. Rats were tested in the tail flick test before and after microinjections of the cannabinoid agonist WIN55, 212-2 (5 microg) into one of 17 different brain regions. WIN55,212-2 significantly elevated tail-flick latencies when injected into the amygdala, the lateral posterior and submedius regions of the thalamus, the superior colliculus and the noradrenergic A5 region. By contrast, pain behavior was unaffected by microinjections of the cannabinoid into the other 11 areas examined (prefrontal cortex, nucleus accumbens, lateral hypothalamus, substantia nigra, cuneiform nucleus, anterior pretectal, intralaminar, parafasicular, posterior, thalamic nuclei, as well as the ventral medial, ventral lateral nuclei in the posterior thalamus).

Suppression of noxious stimulus-evoked activity in the ventral posterolateral nucleus of the thalamus by a cannabinoid agonist: correlation between electrophysiological and antinociceptive effects.

The CNS contains a putative cannabinergic neurotransmitter and an abundance of G-protein-coupled cannabinoid receptors. However, little is known about the function of this novel neurochemical system. Cannabinold agonists produce antinociception in behavioral tests, suggesting the possibility that this system serves in part to modulate pain sensitivity. To explore this possibility, the effects of the cannabinoid agonist WIN 55,212-2 on nociceptive neurons in the ventroposterolateral (VPL) nucleus of the thalamus were examined in urethane-anesthetized rats. After identification of a nociresponsive neuron, a computer-controlled device delivered graded pressure stimuli to the contralateral hindpaw. WIN 55,212-2 (0.0625, 0.125, and 0.25 mg/kg, i.v.) suppressed noxious stimulus-evoked activity of VPL neurons in a dose-dependent and reversible manner. Noxious stimulus-evoked firing was affected more than spontaneous firing. These effects were apparently mediated by cannabinoid receptors, because the cannabinoid receptor-inactive enantiomer of the drug (WIN 55,212-3, 0.25 mg/kg) failed to alter the activity of this population of cells. Administration of morphine (0.5 mg/kg, i.v.) produced effects that were very similar to those produced by the cannabinoid. WIN 55,212-2 (0.25 mg/kg, i.v.) failed to alter the responses of non-nociceptive low-threshold mechanosensitive neurons in the VPL WIN 55,212-2 produced antinociceptive effects with a potency and time course similar to that observed in the electrophysiological experiments, despite the differences in the anesthetic states of the animals used in these experiments. The antinociceptive and electrophysiological effects on VPL neurons outlasted the motor effects of the drug. Furthermore, the changes in nociceptive responding could not be attributed to changes in skin temperature. Taken together, these findings suggest that cannabinoids decrease nociceptive neurotransmission at the level of the thalamus and that one function of endogenous cannabinoids may be to modulate pain sensitivity.

Enhanced cytotoxic potential of alveolar macrophages from cigarette smokers.

Cigarette smoking increases the numbers and oxidative metabolism of alveolar macrophages. Increased production of superoxide (O2-) and H2O2 by alveolar macrophages may contribute to the pathogenesis of cigarette-induced lung diseases. The cytotoxicity mediated by alveolar macrophages from smokers (n = 11) and nonsmokers (n = 13) was compared in an in vitro assay in which the target cells were chromium 51-labeled lung explants. The spontaneous cellular cytotoxicity mediated by smoker macrophages was significantly greater than that of nonsmoker macrophages (cytotoxic index 20.3% +/- 1.9% compared with 5.5% +/- 0.9%, P less than 0.001). Phorbol myristate acetate significantly increased the cytotoxic index of nonsmoker macrophages but did not cause further increases in smoker macrophage killing. The antioxidants superoxide dismutase and catalase produced partial inhibition of smoker macrophage cytotoxicity, suggesting that target cell killing was mediated in part by oxidant mechanisms. Supplementation of smokers' diets with high-dose oral vitamin E failed to decrease smoker alveolar macrophage cytotoxicity. These findings demonstrate that smoker alveolar macrophages possess enhanced cytotoxic potential for normal lung parenchymal cells.

Coagulase-negative staphylococcal bacteremia in patients receiving immunosuppressive therapy.

From January 1977 to June 1980, coagulase-negative staphylococci caused bacteremia in 22 (17%) of 130 patients receiving immunosuppressive therapy and were the most common cause of all bacteremias. Sixteen (73%) of the 22 patients had granulocytopenia, and eight were isolated in a laminar air-flow room. A Broviac or Hickman central intravenous (IV) catheter was present in 20 (91%) of 22 patients, and soft-tissue inflammation at the catheter exit site was a significant risk factor for bacteremia. Except for debilitating fevers and local mucocutaneous infections, there were no distinguishing clinical features in patients with bacteremia. Most infections responded to cefazolin sodium or vancomycin hydrochloride therapy; catheter removal was necessary in only seven patients. These data show that coagulase-negative staphylococci can be important pathogens in patients receiving immunosuppressive therapy, even when the patients are isolated in a laminar air-flow room, if normal mucocutaneous, host-defence barriers are interrupted by IV catheter-insertion or chemotherapy.

Effect of silver nitrate application on the conjunctival flora of the newborn: and the occurrence of clostridial conjunctivitis.

Newborn conjunctival cultures were obtained from 35 babies prior to silver nitrate application and 48 hours later. On initial culture, 46 facultative bacteria and 27 anaerobes were recovered; 48 facultative and 18 anaerobes were recovered after 48 hours. Haemophilus vaginalis, Bacteroides species and anaerobic cocci decreased in numbers, whereas S. epidermidis, Micrococcus and Propionibacterium acnes increased during this time interval. Clostridial species were isolated from two cases who developed conjunctivitis, along with Peptostreptococcus in one of the cases. In vitro experiments demonstrated lack of killing of C. perfringens in silver nitrate concentrations of 0.1 percent, even after 24 hours exposure.

In vitro studies with netilmicin compared with amikacin, gentamicin, and tobramycin.

Netilmicin, a new semisynthetic aminoglycoside derived by ethylation of the 1-N position of the deoxystreptamine ring of sisomicin, was tested in vitro with 4,070 strains of gram-negative bacilli isolated at the UCLA Medical Center during 1975 to 1976, using the agar dilution technique and an inoculum of approximately 10(4) organisms. Results were compared with those simultaneously obtained for amikacin, gentamicin, and tobramycin. Using Mueller-Hinton medium, inhibitory concentrations in broth correlated with those obtained by the agar dilution method except for Pseudomonas aeruginosa, where a 2- to 16-fold difference in susceptibility was noted. For most clinically significant Enterobacteriaceae and P. aeruginosa, the activity of netilmicin in vitro was comparable or superior to that of gentamicin, tobramycin, and amikacin with respect to potency by weight and achievable blood levels. Against gentamicin-resistant strains (MIC > 16 mug/ml), the activity of netilmicin paralleled that of amikacin with the exception of Providencia stuartii, which was inhibited by amikacin but not by netilmicin.

Comparison of media for isolation of salmonellae and shigellae from fecal specimens.

Five transport media, eight plating media, and three enrichment broth media for the isolation of salmonellae and shigellae were evaluated. Eight laboratories in widely separated regions of the United States participated in this evaluation by submitting 490 fecal specimens in the transport media provided. The results suggest that the newer transport media may not offer any advantage over the use of buffered glycerol-saline in the isolation of these enteric pathogens. Shigellae were best isolated by direct inoculation, whereas salmonellae were isolated in greater numbers after tetrathionate (without Brilliant Green) enrichment with subsequent culturing on the plating medium. The use of a variety of plating media is recommended for the recovery of a larger number of these enteric pathogens.