Effects of alosetron on spontaneous migrating motor complexes in murine small and large bowel in vitro.

Alosetron (Lotronex) is a serotonin subtype 3 (5-HT3) receptor antagonist that alleviates symptoms of irritable bowel syndrome (IBS) in female patients. Alosetron may act centrally, involve the alteration of ascending pain sensation, or modulate peristaltic, secretory, or sensory function. To investigate further the mechanisms underlying its action and gender selectivity we recorded the effect of increasing concentrations of alosetron or ondansetron on spontaneous migrating motor complexes (MMCs) from isolated terminal ileum or colon from C57BL/6 mice. Both antagonists inhibited MMC frequency before affects on duration or amplitude. The threshold of inhibition for alosetron was 100-fold less in small intestine from females (20 nM) than from males. The opposite effect of gender was observed with ondansetron in the colon. All MMCs were abolished by either drug at 10 microM. Our results demonstrate that alosetron selectively inhibits MMC frequency in isolated preparations of murine bowel. Because contractile events in the ileum correlate with symptoms of IBS in humans, the gender selectivity of alosetron may be caused by a direct action within the small intestine.

Spontaneous migrating motor complexes occur in both the terminal ileum and colon of the C57BL/6 mouse in vitro.

We have studied migrating motor complexes (MMCs) in the isolated terminal ileum or colon (IMMCs and CMMCs respectively) of the C57BL/6 mouse. Periodic contractions occurred spontaneously in both preparations in the absence of intraluminal stimulation. After an initial period, complexes became synchronized between the oral and anal ends of the tissue, and could be observed for in excess of 7 h. The propagation velocity was 3.1+/-1.0 and 3.9+/-0.6 mm s(-1) in the ileum and colon respectively. IMMCs occurred every 6.01+/-0.39 min and had a duration of 86.3+/-10.4 s. The interval between CMMCs was smaller (3.52+/-0.31 min) and contractions were shorter in duration (30.7+/-3.6 s). In both preparations, these motor events were dependent on cholinergic transmission: blocked by hexamethonium (500 microM) and attenuated or blocked by atropine (1 microM). This study is the first demonstration of spontaneous migrating contractions in the isolated ileum or colon of the C57BL/6 mouse, the strain of choice for neurological transgenic and targeted mice.

Genetically-targeted and conditionally-regulated ablation of astroglial cells in the central, enteric and peripheral nervous systems in adult transgenic mice.

Ablation of tissue regions, specific genes, or specific cell types represent important means of studying function in the nervous system. Here we summarize recent experience using a strategy for the genetically-targeted and conditionally regulated ablation of astroglial cells in different parts of the nervous system. The strategy is based on the targeted expression of herpes simplex virus thymidine kinase to astroglial cells using the glial fibrillary acid protein promoter in transgenic mice, combined with treatment with the antiviral agent ganciclovir. Under different experimental conditions we find that transgene-expressing astroglial cells can be selectively ablated by ganciclovir in the enteric nervous system, or in the injured forebrain or sciatic nerve, providing models in which to study the functions of these cells.

Leukocyte infiltration, neuronal degeneration, and neurite outgrowth after ablation of scar-forming, reactive astrocytes in adult transgenic mice.

Reactive astrocytes adjacent to a forebrain stab injury were selectively ablated in adult mice expressing HSV-TK from the Gfap promoter by treatment with ganciclovir. Injured tissue that was depleted of GFAP-positive astrocytes exhibited (1) a prolonged 25-fold increase in infiltration of CD45-positive leukocytes, including ultrastructurally identified monocytes, macrophages, neutrophils, and lymphocytes, (2) failure of blood-brain barrier (BBB) repair, (3) substantial neuronal degeneration that could be attenuated by chronic glutamate receptor blockade, and (4) a pronounced increase in local neurite outgrowth. These findings show that genetic targeting can be used to ablate scar-forming astrocytes and demonstrate roles for astrocytes in regulating leukocyte trafficking, repairing the BBB, protecting neurons, and restricting nerve fiber growth after injury in the adult central nervous system.

Fulminant jejuno-ileitis following ablation of enteric glia in adult transgenic mice.

To investigate the roles of astroglial cells, we targeted their ablation genetically. Transgenic mice were generated expressing herpes simplex virus thymidine kinase from the mouse glial fibrillary acidic protein (GFAP) promoter. In adult transgenic mice, 2 weeks of subcutaneous treatment with the antiviral agent ganciclovir preferentially ablated transgene-expressing, GFAP-positive glia from the jejunum and ileum, causing a fulminating and fatal jejuno-ileitis. This pathology was independent of bacterial overgrowth and was characterized by increased myeloperoxidase activity, moderate degeneration of myenteric neurons, and intraluminal hemorrhage. These findings demonstrate that enteric glia play an essential role in maintaining the integrity of the bowel and suggest that their loss or dysfunction may contribute to the cellular mechanisms of inflammatory bowel disease.