Particular Strain Of Mice Research Articles

Supraphysiological Levels of IL-2 in Jak3-Deficient Mice Promote Strong Proliferative Responses of Adoptively Transferred Naive CD8+ T Cells.

The antigen-independent, strong proliferative responses of naive CD8+ T cells have been well demonstrated in a particular strain of mice lacking IL-2 receptors. This type of proliferation is mainly driven by common gamma-chain (γc) cytokines, such as IL-2, IL-7, and IL-15, present at abnormally high levels in these mice. Similarly, in the present study, we showed that mice lacking Janus kinase 3 (Jak3), a tyrosine kinase crucial for γc cytokine signaling, could induce strong proliferation of adoptively transferred naive CD8+ T cells. This proliferation was also independent of antigenic stimulation, but heavily dependent on IL-2, as evidenced by the failure of proliferation of adoptively transferred IL-2 receptor alpha- and beta-chain-deficient naive CD8+ T cells. Consistent with this, Jak3 –/– mice showed elevated serum levels of IL-2 compared to wild-type mice, and interestingly, IL-2 production was due to high levels of accumulation of activated CD4+ T cells in Jak3 –/– mice along with defective CD4+ T regulatory cells. Collectively, these findings reveal previously unidentified unique immune contexts of Jak3 –/– mice that cause robust IL-2-driven T cell expansion and have a clinical implication for designing a treatment strategy for human patients with loss-of-function genetic mutations of Jak3.

Pilin Vaccination Stimulates Weak Antibody Responses and Provides No Protection in a C57Bl/6 Murine Model of Acute Clostridium difficile Infection.

Clostridium difficile is the leading cause of nosocomial infections in the United States, adding billions of dollars per year to health care costs. A vaccine targeted against the bacterium would be extremely beneficial in decreasing the morbidity and mortality caused by C. difficile-associated disease; a vaccine directed against a colonization factor would hinder the spread of the bacterium as well as prevent disease. Type IV pili (T4Ps) are extracellular appendages composed of protein monomers called pilins. They are involved in adhesion and colonization in a wide variety of bacteria and archaea, and are putative colonization factors in C. difficile. We hypothesized that vaccinating mice with pilins would lead to generation of anti-pilin antibodies, and would protect against C. difficile challenge. We found that immunizing C57Bl/6 mice with various pilins, whether combined or as individual proteins, led to low anti-pilin antibody titers and no protection upon C. difficile challenge. Passive transfer of anti-pilin antibodies led to high serum anti-pilin IgG titers, but to undetectable fecal anti-pilin IgG titers and did not protect against challenge. The low antibody titers observed in these experiments may be due to the particular strain of mice used. Further experiments, possibly with a different animal model of C. difficile infection, are needed to determine if an anti-T4P vaccine would be protective against C. difficile infection.

Oral ingestion of Capsaicin, the pungent component of chili pepper, enhances a discreet population of macrophages and confers protection from autoimmune diabetes.

Vanilloid receptor 1 (VR1) is expressed on immune cells as well as on sensory neurons. Here we report that VR1 can regulate immunological events in the gut in response to its ligand Capsaicin (CP), a nutritional factor, the pungent component of chili peppers. Oral administration of CP attenuates the proliferation and activation of autoreactive T cells in pancreatic lymph nodes (PLNs) but not other lymph nodes, and protects mice from development of type 1 diabetes (T1D). This is a general phenomenon and not restricted to one particular strain of mice. Engagement of VR1 enhances a discreet population of CD11b(+)/F4/80(+) macrophages in PLN, which express anti-inflammatory factors interleukin (IL)-10 and PD-L1. This population is essential for CP-mediated attenuation of T-cell proliferation in an IL-10-dependent manner. Lack of VR1 expression fails to inhibit proliferation of autoreactive T cells, which is partially reversed in (VR1(+/+) → VR1(-/-)) bone marrow chimeric mice, implying the role of VR1 in crosstalk between neuronal and immunological responses in vivo. These findings imply that endogenous ligands of VR1 can have profound effect on gut-mediated immune tolerance and autoimmunity by influencing the nutrient-immune interactions.

Differential responses to cyclosporine A in a delayed type hypersensitivity model

DTH is classically defined as a model of inflammation involving antigen presentation, T cell activation and cytokine production. Understanding the etiology of the reaction is important, as DTH is commonly used to assess the efficacy of reagents targeting pathways involved in inflammation. We studied the sRBC induced DTH response in two different mouse strains, the commonly used Balb/c and autoimmune prone B6D2F1. Both mouse strains exhibited equivalent paw swelling, but interestingly, CsA treatment reduced swelling in Balb/c but not in B6D2F1 mice. Restimulation of T cells from the draining lymph nodes demonstrates that Balb/c T cells produce greater levels of IFNγ, TNFα and IL-10 compared to B6D2F1 T cells, suggesting that Balb/c DTH may be more cytokine mediated than B6D2F1 DTH. Moreover, IFNγ deficient mice on the Balb/c background have reduced DTH. Although the development of autoantibodies in B6D2F1 mice has been linked to IL-17, we found that treatment with αIL-17A or αIL-17F had no effect on DTH in either strain. Our results indicate that the pathology of the DTH response is different in Balb/c compared to B6D2F1 mice, implying that different etiologies are involved. Therefore, understanding the DTH response in a particular strain of mouse is important in order to accurately determine the specific pathways responsible for the resultant inflammation.

Social defeat stress, sensitization, and intravenous cocaine self-administration in mice

Behavioral sensitization has been proposed as a process that is important in compulsive drug use and in psychotic disorders. The present experiments examine the relationship between behavioral sensitization, induced by either social defeat or amphetamine, and intravenous cocaine self-administration in mice. Male CFW mice were exposed either to defeat experiences, amphetamine (2.5 mg/kg, i.p.) or saline (i.p.) every day for 10 days. Ten days after the last defeat or injection, mice were challenged with varying doses of amphetamine (1.0-2.5 mg/kg i.p). Mice were then trained to nose poke for intravenous cocaine (1.0 mg/kg/inf) during daily 3-h sessions. Following this acquisition phase, the animals self-administered varying doses of cocaine (0.3-1.8 mg/kg/inf) or were allowed to self-administer cocaine (0.3 mg/kg/inf) according to a progressive ratio schedule of reinforcement. Repeated social defeat produced a sensitized motor response to a single challenge of 1.5 mg/kg amphetamine and to a cumulative dosing of amphetamine. Amphetamine-pretreated mice exhibited increased cocaine self-administration during acquisition and elevated break points during performance on a progressive ratio schedule of reinforcement relative to stress-sensitized and control animals. These data extend the evidence from rats to mice for the process of sensitization leading to more cocaine taking. Contrary to what is seen in rats, increased levels of cocaine self-administration were seen only in the amphetamine-pretreated mice and not after repeated defeat stress, suggesting that the sensitized response to defeat stress may not be as robust as it is in rats in this particular strain of mice.

Behavioral and molecular consequences of repeated social defeat stress: sensitization and intravenous cocaine self‐administration in mice

The process of sensitization may be important in the development of drug addiction, and stress exposure can increase cocaine self-administration. We examined the relationship between sensitization induced by social defeat stress and intravenous cocaine self-administration in male CFW mice, and compared this to the cocaine taking of amphetamine-sensitized mice. In addition, we differentiated the stress of being dominant from being defeated by examining Fos expression in raphe cells of mice that experienced 10 daily aggressive encounters. Repeated defeat produced a sensitized motor response to amphetamine, while repeated offensive aggression did not, and this may be related to differential cellular activation in the raphe. Defeated mice showed increased Fos activity in the dorsal raphe, whereas stressed residents exhibited blunted Fos expression in the median raphe following amphetamine challenge. Amphetamine-sensitized mice took more cocaine during acquisition and reached higher break points during performance on a progressive ratio schedule, relative to stress-sensitized and control mice, extending the evidence from rats to mice for the process of sensitization facilitating cocaine taking. Augmented cocaine intake was not seen after repeated defeat, suggesting that the sensitized response resulting from defeat stress may not be enduring or robust in this particular strain of mouse. Support contributed by: DA02623 and DA018478

Mitochondrial production of reactive oxygen species and incidence of age-associated lymphoma in OF1 mice: Effect of alternate-day fasting

There is currently of a great interest investigating the role of nutrition in the prevention of age-associated disorders. The present study aimed to evaluate, on a particular strain of mice, the efficacy of alternate-day fasting on the mitochondrial production of free radical species and on the incidence of a specific cancer (lymphoma) in aged mice. Alternate fasting, that was initiated in middle age mice through a 4 month period, reduced significantly the incidence of lymphoma (0% versus 33% for controls). No remarkable difference was observed in the overall food consumption between alternate-fed (AF) and ad libitum (AL) mice, suggesting that the efficacy of alternate fasting did not really depend on calorie restriction. A significant decrease in the mitochondrial generation of reactive oxygen species (ROS) that was associated with a significant increase in spleen mitochondria SOD activity was observed when mice were maintained on intermittent fasting. Our results suggest that alternate fasting could exert a beneficial antioxidant effect and a modulation of the oxidative stress associated with aging.

Tumor-Free, But Not in the Clear

COLD SPRING HARBOR, NEW YORK --Like a good mother, the p53 protein watches over a cell to protect its DNA. But a deviant partner spurs p53 to sour an organism's future. An altered version of the tumor-blocker accelerates degeneration and makes mice small, apparently by sparking a hormone pathway, according to research presented here 3 October at the Cold Spring Harbor Laboratory's Molecular Genetics of Aging meeting. p53 halts cancer by preventing damaged cells from dividing or by goading them to commit suicide. Previous work suggested that stabilizing p53 accelerates mouse aging despite making cancer infrequent, although questions remain because the animal carries genetic alterations in addition to the one in p53 (see Campisi Perspective ). Part of p53 is missing in a particular strain of mice infected with a leukemia virus. To learn more about the variant, geneticist Heidi Scrable and colleagues at the University of Virginia, Charlottesville, modified mice to carry a gene that produces the shortened version of p53, dubbed p44. Rodents that pump out p44 and p53 developed no tumors but showed signs of premature aging, such as reduced muscle mass and decreased fertility; the animals were also half normal size. p53 molecules work in teams of four; p44 might infiltrate to create mixed groups. Because p44 lacks a region that causes its degradation, the new quartet is probably unusually stable. A drop in the animals' cell number might underlie their size difference: Mothers containing p44 produced embryos that harbored fewer cells than normal, although the cells were regular size. Moreover, cells cultured from p44-carrying embryos proliferated slowly. Scrable speculates that the introduction of p44 prods p53 to slow cell division. The animals' small size hinted that growth-stimulating pathways might be blocked. But further experiments suggest that p44 and p53 together instead crank up the insulin-like growth factor-1 (IGF-1) pathway, a molecular relay that promotes growth and perhaps aging in mammals (see Bartke Viewpoint ); a related pathway curbs life-span in flies and nematodes (see Strauss Science Article ). Compared with controls, cells from the p44-containing animals harbored large amounts of the IGF-1 receptor protein as well as the activated form of Akt, a protein that it stimulates. p53 normally blocks these components of the IGF-1 pathway. Scrable doesn't know why the p44-p53 combination provokes rather than suppresses the pathway, but the effect might stem from the fact that p44 lacks a segment to which regulatory proteins bind and it therefore alters how p53 turns on genes. The IGF-1 changes match observations in nematodes, says molecular geneticist Cynthia Kenyon of the University of California, San Francisco. "It's really clear that if you turn up IGF-1[-related signaling], worms have a short life-span." What isn't clear is why the animals are small, says Kenyon. Perhaps another altered cellular signal overrides IGF-1's growth-spurring effect, she suggests. The study backs the emerging hypothesis that unchecked p53 causes premature aging, says geneticist Ronald dePinho of Harvard University, and it is the first to suggest that p53 acts early in embryonic development. Further work could reveal how to help p53 raise a happy cellular family. --R. John Davenport W. Gluba, A. Sutherland, H. Scrable, Growth deficiency and early aging in p44/p53 mice. Cold Spring Harbor Laboratory, Molecular Genetics of Aging, 2 to 6 October 2002, Cold Spring Harbor, New York. [Meeting Home Page]

A cascade of cytokines in toxoplasmosis

A pair of papers describing a genetic defect in the interleukin (IL)-12 signaling system of a particular strain of mice has interesting relevance to toxoplasmosis. An in vitro analysis of the defect itself is presented by Ortmann, R. et al. (2001) (J Immunol. 166, 5712–5719). George Yap et al. (2001) (J. Immunol. 166, 5720–5725) use the same mice (a substrain of B10.Q/J) to demonstrate a crucial role for IL-12 signaling in mediating acute resistance to Toxoplasma gondii infection. In spite of robust systemic production of IL-12, the mice fail to mount an early interferon (IFN)-γ response after parasite inoculation. It is not until later, when the infection has already taken hold, that a delayed IL-12-mediated IFN-γ response emerges. The early IFN-γ deficiency and host resistance can be corrected by administration of IL-18, suggesting a crucial role for a cascade of IL-12, IL-18 and IFN-γ production for early resolution of toxoplasma infection. SHK

Behavioral impairment in radial-arm maze learning and acetylcholine content of the hippocampus and cerebral cortex in aged mice

Age-related changes in spatial learning performance were studied in relation to acetylcholine (ACh) content of brain regions in male aged (28-month-old) and young (5-month-old) mice of BDF1 strain. As there were large individual differences in the spatial performance of aged mice, the aged mice were divided into two subgroups, old A and old B. The old A group included the six best performers out of the 12 aged mice and the old B group included the remaining 6 worst performers. In a radial-arm maze task with 8 baited arms, aged mice in the old B group showed a marked deficit in acquisition performance and habituation to the apparatus. In the more difficult maze task with only 4 baited arms, the aged mice in the old B group exhibited marked impairment both in working memory and reference memory throughout training, whereas the aged mice in the old A group showed deficits in reference memory during the first 20 days of training and working memory during the last 20 days relative to young mice. Neurochemical analysis revealed significant decreases in the ACh content of the hippocampus and striatum in both aged groups, and in the frontal cortex and posterior cortex of the old B group as compared to the young group. Correlational analysis showed significant correlations between learning performance in the spatial task and ACh levels in the hippocampus, frontal cortex, and posterior cortex. The present results demonstrate that a marked decrease in ACh levels in the hippocampus and cerebral cortex may be closely related to behavioral impairment in spatial learning and habituation in fully aged mice. Thus, this particular strain of mice may serve as a useful animal model for age-related impairment of cognitive functions in relation to hypofunction of the cholinergic systems in the basal forebrain.

L'action du génome paternel au début du développement embryonnaire

Recent experiments using nuclear transfer between mouse embryos shows that the parental genomes are not equivalent: both of them must be present at least in a part of the blastomeres to allow full term development of the embryo. The paternal genome is preferentially involved in the development of trophoectodermic cells and appears to be eliminated from the primitive ectoderm when kept alone in androgenetic cells aggregated with normal ones. Differences of expression between parental genomes affect not only the X chromosome, but also several autosomes as evidenced by genetic analysis using meiotic non-disjunction. The nature of the imprinting mechanism that leads to the modification of parental homologous chromosomes is unknown. From experiments involving a particular strain of mice called DDK it appears that the egg cytoplasm may exert a specific action not only on the male pronucleus at the one-cell stage, but also on the paternal contribution of the diploid nucleus of the embryo at the 2-cell stage; a paternal developmental effect on the cytoplasm of one-cell stage embryo can also be demonstrated. These results show that nucleocytoplasmic interactions regulate early paternal gene expression differently from the maternal one.

Stressor-provoked behavioral changes in six strains of mice.

Behavioral changes induced by inescapable shock were examined in six strains of mice. Exposure to shock provoked time-dependent disturbances of shuttle escape performance. In some strains the shock treatment did not affect escape performance, whereas in others profound performance deficits were evident. The inescapable shock treatment likewise induced strain-dependent alterations of performance in a forced-swim task. In most instances the shock treatment initially provoked invigorated responding, but in other strains the shock had no effect or depressed active responding. Finally, Y-maze spontaneous alteration performance was not affected by the shock treatment, although a strain-dependent increase of perseverative responses was evident. The occurrence of a stressor-induced deficit in one task in a particular strain of mouse was not predictive of behavioral alterations in a second task. These data are discussed with respect to animal models of depression and genetic differences associated with the response to stressors.

Activation of a cellular proto-oncogene in spontaneous liver tumor tissue of the B6C3F1 mouse.

The controversy surrounding the interpretation of observed increases in the high spontaneous liver tumor incidence of the B6C3F1 mouse after administration of certain chemical agents necessitates a mechanistic understanding into the nature of tumor development in this particular strain of mouse. Recently, cancer genes (oncogenes) have been detected in the DNA from a variety of human tumors and tumor cell lines. These genes have been implicated to play a role in the transformation of normal cells into cancerous ones. To investigate the role that cellular oncogenes might play in the development of spontaneous liver tumors in the B6C3F1 mouse, DNA was isolated from spontaneously occurring liver tumors and transfected into NIH 3T3 fibroblasts. DNA from this tumor tissue was capable of transforming NIH 3T3 cells from 82% of the animals examined strongly suggesting the presence of an active cellular oncogene. In contrast, DNA isolated from surrounding non-tumorous liver tissue and liver tissue from non-tumor bearing mice did not cause any transformation in the NIH 3T3 assay. These data demonstrate that the active cellular oncogene is not present in the hepatic tissue via a germ-line transmission but is activated only in those cells of the tumor tissue. Experiments using Southern blot hybridization analysis have identified this active cellular oncogene to be a member of the ras oncogene family. Identification of this cellular oncogene will now allow the evaluation of factors which might modify its expression. These future studies will lead to an increased understanding of potential mechanisms by which hepatic tumors are enhanced and should provide more informed estimates of risk for man based on bioassay data generated in this strain of mouse.

Paradoxical blockade of a muscimol response by thip, a GABA agonist and also by GABA-transaminase inhibitors

In a particular strain of mice, relatively large doses of muscimol caused myoclonic jerks of high frequency. This muscimol response was blocked in a dose-dependent manner by a γ-aminobutyric acid (GABA) agonist 4,5,6,7-tetrahydroisoxazolo(5,4-c)pyridin-3-ol-hydrate (THIP). Studies using γ-acetylenic GABA and γ-vinyl GABA demonstrated that the blockade of the muscimol-induced jerks caused by these irreversible GABA-transaminase inhibitors closely paralleled the elevation of brain GABA level. It seems that, in this particular strain of mice, muscimol, or one of its metabolites, possibly acts on certain specific binding sites in the central nervous system, eliciting myoclonic jerks and that these receptor sites are different from those to which GABA or THIP binds. Earlier studies demonstrated the value of the muscimol-induced myoclonic jerks as an animal model for postanoxic action myoclonus. Based on this finding, it is proposed that both THIP and the GABA-transaminase inhibitors might prove to be of value in the management of this clinical condition.

Antimyoclonic Effect of Sodium Oxybate: Clinical Implications

To the Editor.— In several neurological disorders, myoclonus appears as a disturbing and persistent symptom. Presently, these involuntary and repetitive movements of the skeletal muscle are being controlled by the administration of the serotonin precursor carubicin hydrochloride in combination with a peripheral decarboxylase inhibitor1or by the benzodiazepine muscle relaxant clonazepam.2While these two drug treatments have been found to be effective, their long-term administration could lead to undesirable side effects. Dependence liability of the benzodiazepines is well documented. Because of the nonavailability of a suitable animal model for myoclonus in the past, introduction of new drugs for the management of clinical myoclonus was not possible. Recently we introduced a valid animal model for clinical myoclonus.3This is based on our observation that intraperitoneal (IP) administration of muscimol, 3 mg/kg, a γ-aminobutyric acid agonist, induces involuntary and repetitive myoclonic jerks in a particular strain of mice that

Demonstration of the antimyoclonic effect of 1-hydroxy-3-amino-pyrrolidone-2 (HA-966) using a new animal model

Comparatively high doses of muscimol induces myoclonic jerks in a particular strain of mice. 1-Hydroxy-3-amino-pyrrolidone-2 (HA-966) in doses of 1–10 mg/kg caused a dose-dependent blockade of this response of muscimol. Our earlier studies showed that the muscimol-induced myoclinic jerks fulfill certain criteria necessary to be considered as a valid animal model for postanoxic action myoclonus. On this basis it was concluded that HA-966 might prove to be of value in the management of this disorder.

Induction of cytolytic T- and B-cell responses against influenza virus infections.

Inoculation of mice with live influenza virus results in the induction of cytotoxic thymus-derived (T) lymphocytes and of bone marrow-derived (B) cells producing antiviral antibodies. An assay system was developed to evaluate both types of immune responses on a cellular basis within the same lymphocyte pool with no need to separate out the different effector cell classes. The test system represented a modification of the 51Cr-release assay. T-cell activity was measured exclusively in the absence of active complement using targets that were compatible for determinants encoded by the mouse major histocompatibility gene complex, H-2. H-2-different and even xenogeneic target cells were lysed in the presence of either non-inactivated fetal calf serum or normal rabbit serum as a complement source. Cytotoxicity was mediated in the latter case by direct interaction of B-cell-produced immunoglobulin directed to viral antigens expressed by the target cell and complement. Antibody-dependent cell-mediated cytotoxicity mechanisms did not contribute to cytotoxicity in the test system described. It was demontrated that the cytolytic B-cell responses of one particular strain of mice (BALB/c) against different influenza A viruses were restricted to the immunizing virus on the effector cell level. In another strain of mice (C3H), B cells revealed a broad cross-reactive response resembling that of killer T cells.

Susceptibility of Five Strains of Mice to Babesia microti of Human Origin

One outbred (CF1) and four inbred (BALB/c, C57, CBA and C3H) strains of mice were tested for susceptibility to Babesia microti of human origin. Of these, intact C3H mice developed higher parasitemia than all other intact mice, while BALB/c mice developed the highest parasitemia among splenectomized mice. Susceptibility was not related to H-2 haplotype in any obvious way. Because C3H and BALB/c mice developed relatively high initial peak parasitemias, the parasite was serially passaged in both of these mouse strains in an attempt to increase parasite virulence. After 30 passages in BALB/c and 49 passages in C3H mice over a period of 12 months, maximum parasitemias were 50 times higher than those observed initially. After the peak parasitemias of these two mouse-adapted parasites had stabilized, the relationship between onset and level of maximum parasitemia and number of parasites inoculated was determined. With both C3H- and BALB/c-adapted parasites, as inoculum size increased, the time required to reach maximum parasitemia decreased and the level of maximum parasitemia increased. Studies involving infection of either mouse strain with parasites adapted to the heterologous mouse strain indicated that C3H mice were more susceptible than BALB/c mice to homologous or heterologous parasites. These data suggest that the virulence of B. microti to the mouse can be increased by prolonged passage in this host. Once adaptation to this host species has occurred, virulence appears to be more dependent on the innate susceptibility of the mouse strain than on adaptation of the parasites to a particular strain of mouse.

Comparison of the effects of anisomycin on memory across six strains of mice

Retention for step-through passive avoidance training was compared across six strains of mice including pigmented and albinos, inbred and outbred, and strains light and heavy in weight. Widely differing conditions of training were required in order for 90–100% of the mice in each strain to learn one-trial passive avoidance. The effect of anisomycin on protein synthesis was investigated and no major differences were found across strains. A single injection of anisomycin, given 15 min prior to training, was found to cause significant amnesia for passive avoidance training in all strains of mice. It was also found that a small increase in shock intensity decreased the percent amnesia caused by a single pretraining injection of anisomycin. If at this higher shock intensity the pretraining injection was followed by a second injection of anisomycin, given 1.75 hr after training, a high percentage of the subjects was again found to have amnesia. Discussion includes (a) possible causes of strain differences in ability to acquire the passive avoidance habit, (b) a general methodology for obtaining amnestic effects independent of the particular strain of mice used, and (c) the necessity of protein synthesis for memory formation.

EFFECT OF SPLENECTOMY ON THE SUSCEPTIBILITY OF MICE INOCULATED WITH DIPLOCOCCUS PNEUMONIAE

An experimental model is described which demonstrated increased susceptibility of mice to infection with D. pneumoniae following splenectomy. It was necessary to use small numbers of a particular strain of pneumococcus (D. pneumoniae type 6), intravenous infection and a particular strain of mouse (pathogen-free NCS strain). The increase in susceptibility persisted for at least 4 months after splenectomy. With modifications in experimental design such as use of large numbers of organisms, a different strain of pneumococcus, the intraperitoneal route of infection or a different mouse strain no increase or a much less impressive increase in susceptibility was demonstrated. Following intravenous injection of small numbers of D. pneumoniae Type 6 bacteremia tended to persist in all NCS mice. Multiplication of pneumococci subsequently occurred in a higher proportion of mice with splenectomy and at a more rapid rate than in control animals. Mice with splenectomy usually had more D. pneumoniae per ml of blood than per gram of any tissue. This suggested that in these mice multiplication of microorganisms occurs primarily in blood. In control mice higher concentrations of bacteria were present in spleen than in blood, and higher concentrations were found in blood than in other tissues. These results suggested that in normal mice infected intravenously with small numbers of D. pneumoniae Type 6, the spleen protects by removing and killing small but critical numbers of D. pneumoniae which are circulating in the blood. No evidence was found to suggest that the altered susceptibility is mediated by an effect of splenectomy on numbers of circulating leukocytes or on the antibacterial activity of mouse blood.