Naloxone and ouabain in ultralow concentrations restore Na+/K+-ATPase and cytoskeleton in lipopolysaccharide-treated astrocytes.

Astrocytes respond to inflammatory stimuli and may be important modulators of the inflammatory response in the nervous system. This study aimed first to assess how astrocytes in primary culture behave in response to inflammatory stimuli concerning intracellular Ca(2+) responses, expression of Toll-like receptor 4 (TLR4), Na(+)/K(+)-ATPase, actin filament organization, and expression of cytokines. In a cell culture model with lipopolysaccharide (LPS), astrocyte response was assessed first in the acute phase and then after incubation with LPS for 1-48 h. The concentration curve for LPS-stimulated Ca(2+) responses was bell-shaped, and the astrocytes expressed TLR4, which detects LPS and evokes intracellular Ca(2+) transients. After a long incubation with LPS, TLR4 was up-regulated, LPS-evoked Ca(2+) transients were expressed as oscillations, Na(+)/K(+)-ATPase was down-regulated, and the actin filaments were disorganized. Interleukin-1ß (IL-1ß) release was increased after 24 h in LPS. A second aim was to try to restore the LPS-induced changes in astrocytes with substances that may have dose-dependent anti-inflammatory properties. Naloxone and ouabain were tested separately in ultralow or high concentrations. Both substances evoked intracellular Ca(2+) transients for all of the concentrations from 10(-15) up to 10(-4) M. Neither substance blocked the TLR4-evoked Ca(2+) responses. Naloxone and ouabain prevented the LPS-induced down-regulation of Na(+)/K(+)-ATPase and restored the actin filaments. Ouabain, in addition, reduced the IL-1ß release from reactive astrocytes. Notably, ultralow concentrations (10(-12) M) of naloxone and ouabain showed these qualities. Ouabain seems to be more potent in these effects of the two tested substances.

A pilot study of colonic B cell pattern in irritable bowel syndrome.

OBJECTIVE: Low-grade gastrointestinal inflammation has been reported in patients with irritable bowel syndrome (IBS). However, the colonic B-cell pattern has not been investigated in these patients. Therefore, the aim of this pilot study was to investigate the distribution and isotype of immunoglobulin-producing B cells in the colonic mucosa of IBS patients. MATERIAL AND METHODS: Patients with IBS (n=12) fulfilling the Rome II criteria were compared with controls (n=11). Immunohistochemical staining of biopsies from the sigmoid and ascending colon was performed. RESULTS: The number of IgA(+) B cells in the ascending colon was lower in IBS patients than in controls (p=0.039). Furthermore, unlike controls, IBS patients had a reduction of IgA(+) B cells in the ascending colon relative to the sigmoid colon (p=0.04). Neither the IgG(+), nor the IgM(+) colonic B-cell numbers differed between IBS patients and controls. Very few colonic IgE(+) cells were detected and there was no difference between the two subject groups. CONCLUSIONS: The reduced number of colonic IgA(+) B cells in IBS patients suggests that the disorder may be associated with a modified gut immune defence. Whether this phenomenon is causally related to symptoms remains unknown and merits further investigation in a larger group of patients.

Anti-inflammatory substances can influence some glial cell types but not others.

In rat microglial enriched cultures, expressing Toll-like receptor 4, we studied cytokine release after exposure with 1 ng/ml LPS for 0.5-24 h. Dexamethasone and corticosterone exposure served as controls. We focused on whether naloxone, ouabain, and bupivacaine, all agents with reported anti-inflammatory effects on astrocytes, could affect the release of TNF-α and IL-1ß in microglia. Our results show that neither ultralow (10(-12) M) nor high (10(-6) M) concentrations of these agents had demonstrable effects on cytokine release in microglia. The results indicate that anti-inflammatory substances exert specific influences on different glial cell types. Astrocytes seem to be functional targets for anti-inflammatory substances while microglia respond directly to inflammatory stimuli and are thus more sensitive to anti-inflammatory substances like corticoids. The physiological relevance might be that astrocyte dysfunction influences neuronal signalling both due to direct disturbance of astrocyte functions and in the communication within the astrocyte networks. When the signalling between astrocytes is working, then microglia produce less pro-inflammatory cytokines.