The commensal microbiota protects the murine host from enteric pathogens. Nevertheless, specific pathogens are able to colonize the intestinal tract and invade, despite the presence of an intact biota. Possibly, effective pathogens disrupt the indigenous microbiota, either directly through pathogen-commensal interaction, indirectly via the host mucosal immune response to the pathogen, or by a combination of these factors. This study investigates the effect of peroral Salmonella enterica serovar Typhimurium infection on the intestinal microbiota. Since the majority of the intestinal microbiota cannot be cultured by conventional techniques, molecular approaches using 16S rRNA sequences were applied. Several major bacterial groups were assayed using quantitative PCR. Administration of either the 50% lethal dose (LD 50 ) or 10؋ LD 50 of Salmonella enterica serovar Typhimurium caused changes in the microbiota throughout the intestinal tract over the time course of infection. A 95% decrease in total bacterial number was noted in the cecum and large intestine with 10؋ LD 50 S. enterica serovar Typhimurium challenge at 7 days postinfection, concurrent with gross evidence of diarrhea. In addition, alterations in microbiota composition preceded the onset of diarrhea, suggesting the involvement of pathogen-commensal interactions and/or host responses unrelated to diarrhea. Microbiota alterations were not permanent and reverted to the microbiota of uninfected mice by 1 month postinfection. Infection with a Salmonella pathogenicity island 1 (SPI1) mutant did not result in microbiota alterations, while SPI2 mutant infections triggered partial changes. Neither mutant was capable of prolonged colonization or induction of mucosal inflammation. These data suggest that several Salmonella virulence factors, particularly those involved in the local mucosal host response, are required for disruption of the intestinal ecosystem.

Roles of endogenous opioid peptides and their receptors in modulation of the nocifensive responses to formalin in mice were studied. Mice were pretreated i.c.v. or intrathecally (i.t.) with selective opioid receptor antagonists or intrathecally with antisera against endogenous opioid peptides and the nocifensive licking responses to intraplantar injection of formalin (0.5%, 25 l) were then observed. Pretreatment with the ⑀-opioid receptor antagonist ␤-endorphin(1-27) or the selective -opioid receptor antagonist D-Phe-Cys-Tyr-Orn-Thr-Pen-Thr-NH 2 (CTOP) given i.c.v. dose dependently enhanced the second, but not the first phase of the nocifensive response. However, i.c.v. pretreatment with the selective ␦-receptor antagonist naltrindole or -receptor antagonist nor-binaltrophimine did not affect the nocifensive responses. Intrathecal pretreatment with selective ␦ 1 -opioid antagonist 7-benzylidene naltrexamine significantly enhanced both the first and second phases of nocifension. Intrathecal pretreatment with CTOP also increased the second but not the first phase of the nocifension. However, i.t. pretreatment with the selective ␦ 2 -receptor antagonist naltriben or nor-binaltrophimine did not affect the second phase of the nocifension. Intrathecal pretreatment with antiserum against Leu-enkephalin, Met-enkephalin, or dynorphin A(1-17), but not ␤-endorphin, enhanced only the second phase of nocifensive response to formalin. It is concluded that the blockade of ⑀-and -receptors, but not ␦-or -receptors, at the supraspinal sites enhanced the second phase of formalin-induced nocifension. In the spinal cord, Leu-enkephalin, and to a lesser extent, Met-enkephalin and dynorphin A(1-17) andand ␦ 1 -opioid receptors, but not ␦ 2 -or -opioid receptors, are involved in modulating the feedback inhibition of the second phase of formalin-induced nocifension.

An unbiased conditioned place preference (CPP) paradigm was used to evaluate the reward effects of endogenous -opioid receptor ligands endomorphin-1 (EM-1) and endomorphin-2 (EM-2) from the mesolimbic posterior nucleus accumbens (Acb) shell and the ventral tegmental area (VTA) in CD rats. EM-1 (1.6 -8.1 nmol) microinjected into posterior Acb shell produced CPP, whereas EM-2 (8.7-17.5 nmol) given into the same Acb shell produced conditioned place aversion (CPA). EM-1 (1.6 -16.3 nmol) microinjected into the VTA produced CPP, whereas EM-2 (8.7 and 17.5 nmol) given into the same VTA site did not produce any effect, but at a high dose (35 nmol) produced CPP. EM-1 (3.3 nmol) or EM-2 (17.5 nmol) microinjected into the nigrostriatal substantia nigra was not significantly different from vehicle-injected groups. D-Phe-Cys-Tyr-D-Trp-Orn-Thr-PenThr-NH 2 (CTOP) at 94.13 pmol or 3-methoxynaltrexone at 0.64 pmol microinjected into the posterior Acb shell blocked EM-1-induced CPP and EM-2-induced CPA. At a higher dose, CTOP (941.3 pmol) and 3-methoxynaltrexone (6.4 pmol) produced CPA and CPP, respectively. Coadministration with antiserum against dynorphin A(1-17) (Dyn) (10 g) microinjected into the posterior Acb shell blocked EM-2-induced CPA. However, it did not affect EM-1-induced CPP. It is concluded that EM-1 and EM-2 produce site-dependent CPP and CPA, respectively, by stimulation of different subtypes of -opioid-receptors; stimulation of one subtype of -opioid-receptor at the posterior Acb shell and VTA by EM-1 induces CPP, whereas stimulation of another subtype of -opioid receptor at the posterior Acb shell, but not the VTA, by EM-2 induces the release of Dyn to produce CPA.

Abstract. The involvement of spinopetal noradrenergic and serotonergic systems in antinociception induced by endomorphin-1 (EM-1) and endomorphin-2 (EM-2) given supraspinally or spinally were investigated in male CD-1 mice. Groups of mice were pretreated intrathecally (i.t.) with 6-hydroxydopamine (6-OHDA, 20 mg) or 5,7-dihydroxytryptamine (5,7-DHT, 50 m g) for 3 days before intracerebroventricular (i.c.v.) or i.t. injection of different doses of EM-1 or EM-2, and the tail-flick response was measured for antinociceptive effects. I.t. pretreatment with 6-OHDA for 3 days, which markedly depleted noradrenaline (NA) contents by more than 90%, but not serotonin (5-HT) in the spinal cord, completely abolished the antinociception induced by i.c.v.-administered EM-1 or EM-2. Intrathecal pretreatment with 5,7-DHT for 3 days, which markedly reduced 5-HT contents by more than 92%, but only reduced NA by 14 -25% in the spinal cord, also markedly attenuated the antinociception induced by i.c.v.-administered EM-1 or EM-2. However, the antinociception induced by i.t.-administered EM-1 or EM-2 was not affected in either 6-OHDA or 5,7-DHT pretreated mice. It is concluded that NA and 5-HT in the spinal cord are involved in the antinociception induced by supraspinally, but not spinally administered EM-1 and EM-2.

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