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Mammalian [beta]-globin loci contain multiple [beta]-like genes that are expressed at different times during development. The murine [beta]-globin locus contains two genes expressed during the embryo stage, Ey and/[beta]h1, and two genes expressed at both the fetal and postnatal stages,[beta]-major and [beta]-minor. Studies of transgenic human [beta]-like globin loci in mice have suggested that expression of one gene at the locus will suppress expression of other genes at the locus. To test this hypothesis we produced mouse lines with deletions of either the Ey or [beta]h1 promoter in the endogenous murine [beta]-globin locus. Promoter deletion eliminated expression of the mutant gene but did not affect expression of the remaining embryonic gene or the fetal/adult [beta]-globin genes on the mutant allele. These results demonstrate a lack of competitive effects between individual mouse embryonic [beta]-globin gene promoters and other genes in the locus. The implication of these findings for models of [beta]-globin gene expression are discussed.

Proliferation and carcinogenesis of the large intestinal epithelial cells (IEC) cells is significantly increased in transgenic mice that overexpress the precursor progastrin (PG) peptide. It is not known if the in vivo growth effects of PG on IEC cells are mediated directly or indirectly. Full-length recombinant human PG (rhP[G.sub.1-80]) was generated to examine possible direct effects of PG on IEC cells. Surprisingly, rhPG (0.1-1.0 nM) was more effective than the completely processed gastrin 17 (G17) peptide as a growth factor. Even though IEC cells did not express CC[K.sub.1] and CC[K.sub.2] receptors (-R), fluorescently labeled G17 and Gly-extended G17 (G-Gly) were specifically bound to the cells, suggesting the presence of binding proteins other than CC[K.sub.1]-R and CC[K.sub.2]-R on IEC cells. High-affinity ([K.sub.d] = 0.5-1.0 nM) binding sites for [sup.I]-rhPG were discovered on IEC cells that demonstrated relative binding affinity for gastrinlike peptides in the order PG [greater than or equal to] COOH-terminally extended G17 [greater than or equal to] G-Gly > G17 > *CCK-8 (* significant difference; P < 0.05). In conclusion, our studies demonstrate for the first time direct growth effects of the full-length precursor peptide on IEC cells in vitro that are apparently mediated by the high-affinity PG binding sites that were discovered on these cells. progastrin-preferring receptors; confocal microscopy; in vitro growth effects; Gly-gastrin

Networks of [gamma]-aminobutyric acid (GABA)ergic interneurons connected via electrical and chemical synapses are thought to play an important role in detecting and promoting synchronous activity in the cerebral cortex. Although the properties of electrical and chemical synaptic interactions among inhibitory interneurons are critical for their function as a network, they have only been studied systematically in juvenile animals. Here, we have used transgenic mice expressing the enhanced green fluorescent protein in cells containing parvalbumin (PV) to study the synaptic connectivity among fast-spiking (FS) cells in slices from adult animals (2-7 months old). We have recorded from pairs of PV-FS cells and found that the majority of them were electrically coupled (61%, 14 of 23 pairs). In addition, 78% of the pairs were connected via GABAergic chemical synapses, often reciprocally. The average coupling coefficient for step injections was 1.5% (n = 14), a smaller value than that reported in juvenile animals. GABA-mediated inhibitory postsynaptic currents and potentials decayed with exponential time constants of 2.6 and 5.9 ms, respectively, and exhibited paired-pulse depression (50-ms interval). The inhibitory synaptic responses in the adult were faster than those observed in young animals. Our results indicate that PV-FS cells are highly interconnected in the adult cerebral cortex by both electrical and chemical synapses, establishing networks that can have important implications for coordinating activity in cortical circuits.

Phenotypic knockout of nerve growth factor (NGF) activity in transgenic anti-NGF mice (AD11 mice) results in a progressive neurodegenerative phenotype resembling Alzheimer's disease. In this article, we examine whether and how the progressive neurodegenerative phenotype of AD11 mice could be prevented or ameliorated by pharmacological treatments with NGF or the cholinergic agonist galantamine, at a relatively early phase of Alzheimer's disease-like neurodegeneration. We demonstrate that the neurodegeneration induced by the expression of anti-NGF antibodies in AD11 mice can be largely reversed by NGF delivery through an olfactory route.

The serine-threonine kinase Akt seems to be central in mediating stimuli from different classes of receptors. In fact, both IGF-1 and IL6-like cytokines induce hypertrophic and antiapoptotic signals in cardiomyocytes through PI3K-dependent Akt activation. More recently, it was shown that Akt is involved also in the hypertrophic and antiapoptotic effects of [beta]-adrenergic stimulation. Thus, to determine the effects of Akt on cardiac function in vivo, we generated a model of cardiac-specific Akt overexpression in mice. Transgenic mice were generated by using the E40K, constitutively active mutant of Akt linked to the rat [alpha]-myosin heavy chain promoter. The effects of cardiac-selective Akt overexpression were studied by echocardiography, cardiac catheterization, histological and biochemical techniques. We found that Akt overexpression produced cardiac hypertrophy at the molecular and histological levels, with a significant increase in cardiomyocyte cell size and concentric LV hypertrophy. Akt-transgenic mice also showed a remarkable increase in cardiac contractility compared with wild-type controls as demonstrated by the analysis of left ventricular ([dP/dt.sub.max]) in an invasive hemodynamic study, although with graded dobutamine infusion, the maximum response was not different from that in controls. Diastolic function, evaluated by left ventricular [dP/dt.sub.min], was not affected at rest but was impaired during graded dobutamine infusion. Isoproterenol-induced cAMP levels, [beta]-adrenergic receptor ([beta]-AR) density, and [beta]-AR affinity were not altered compared with control mice. Moreover, studies on signaling pathway activation from myocardial extracts demonstrated that glycogen synthase kinase3-[beta] is phosphorylated, whereas p42/44 mitogen--activated protein kinases is not, indicating that Akt induces hypertrophy in vivo by activating the glycogen synthase kinase3-[beta]/GATA 4 pathway. In summary, our results not only demonstrate that Akt regulates cardiomyocyte cell size in vivo, but, importantly, show that Akt modulates cardiac contractility in vivo without directly affecting [beta]-AR signaling capacity.

The adult myocardium responds to a variety of pathologic stimuli by hypertrophic growth that frequently progresses to heart failure. The calcium/calmodulin-dependent protein phosphatase calcineurin is a potent transducer of hypertrophic stimuli. Calcineurin dephosphorylates members of the nuclear factor of activated T cell (NFAT) family of transcription factors, which results in their translocation to the nucleus and activation of calcium-dependent genes. Glycogen synthase kinase-3 (GSK-3) phosphorylates NFAT proteins and antagonizes the actions of calcineurin by stimulating NFAT nuclear export. To determine whether activated GSK-3 can act as an antagonist of hypertrophic signaling in the adult heart in vivo, we generated transgenic mice that express a constitutively active form of GSK-3[beta] under control of a cardiac-specific promoter. These mice were physiologically normal under nonstressed conditions, but their ability to mount a hypertrophic response to calcineurin activation was severely impaired. Similarly, cardiac-specific expression of activated GSK-3[beta] diminished hypertrophy in response to chronic [beta]-adrenergic stimulation and pressure overload. These findings reveal a role for GSK-3[beta] as an inhibitor of hypertrophic signaling in the intact myocardium and suggest that elevation of cardiac GSK-3[beta] activity may provide clinical benefit in the treatment of pathologic hypertrophy and heart failure.

Transgenic mice expressing platelet-derived growth factor A chain (PDGF-A) in the distal lung epithelium from the surfactant protein C (SPC) promoter were generated to investigate the role of this growth factor in lung development. Expression of the SPC-PDGFA transgene resulted in an enlarged, nonfunctional lung and perinatal lethality caused by failure to initiate ventilation. Histologic analysis of embryonic day (E) 16.5 lungs revealed increased mesenchymal cells and acinar buds and decreased bronchioles and dilated airspaces in SPC-PDGFA transgenic mice. At E18.5, nontransgenic lungs exhibited lung morphology typical of the saccular stage of lung development, including dilated airspaces, thin respiratory epithelium and mesenchyme, and elastin fiber deposition in primary septa. In contrast, E18.5 transgenic lungs retained many features of the canalicular stage of lung development, including undilated airspaces, cuboidal respiratory epithelium, thickened mesenchyme, and lack of parenchymal elastin deposition. These results indicate that PDGF-A is a potent growth factor for mesenchymal cells in the developing lung and that the downregulation of PDGF-A expression that normally occurs in the lung during late gestation is required for transition from the canalicular to the saccular stage of lung development.

Mice that express hypoxia-inducible factor-1alpha (HIF-1alpha)(Delta)ODD developed a microvasculature with biology different from that described before for either vascular endothelial growth factor (VEGF) or angiopoietin-1 and -2. Hypervascularity induced by HIF-1alpha could improve tissue ischemia therapy.

Functional inactivation of the IGF-I and insulin receptors (IGF-IR and IR) in striated muscle has been found to cause type 2 diabetes. A transgenic mouse with a dominant-negative insulin-like growth factor-1 receptor specifically targeted to the striated muscle was developed for study. The mice, MKR mice, were excellent as a model for this type of study. They showed striking loss of function of the IR and the IGF-IR

The lac operator-repressor system has been found to be functional in mice by transferring an entirely functional lac system to mice. The lac system brings the time dimension of mammalian gene expression in animals under reliable, predictable experimental control. It is likely possible to put mutations, even letahl ones, into the mouse genome and to study them at the organismal level.

Polyglutamine diseases, inherited neurodegenerative deseases, come from a mutational mechanism altering the sequence of a protein so that glutamines are added to a polyglutamine tract in the androgen receptor via one of two possible mechanisms discussed in this review article with information on the effects of polyglutamine expansion at the protein level. Evidence that a polyglutamine tract can itself be toxic exists. Another mechanism would depend on change in the function of the full-length protein. Lessons from Huntington's, and from spinocerebellar ataxia types 1 and 2 are discussed.

Neurofibromatosis type 1 (NF1) has been studied with the finding that ablation of NF1 function in neurons induces abnormal development of the cerebral cortex. It also induces reactive gliosis in the brain. The embryonic lethality of NF1 null mouse embryos was a problem, but by generating a conditional mutation in the NF1 gene it was circumvented. Cre/loxP technology was used. Results show NF1-deficient neurons are able to induce reactive astrogliosis through a non-cell autonomous mechanism and suggest NF1 has a required role in an aspect of CNS development.

Fra-1 has been found to replace c-Fos-dependent functions in mice. Studies with knock out and transgenic mice and structure-function analysis have been used to show two distinct functions for cFos and Fra1, components of the transcription factor activator protein-1 (AP-1). It was seen that Fra-q and c-Fos have kept functional equivalence in vertebrate evolution.

MMP-9 from cells from bone marrow has been found to contribute to carcinogenesis in skin. The matrix metalloproteinase MMP-9/gelatinase B has been found to be upregulated in angiogenic dysplasias and invasive cancers of the epidermis in mice used to study multistage tumorigenesis brought on by HPV1 oncogenes. Transgenic mice without MMP-9 have lower keratinocyte hyperproliferation at all neoplastic stages and lower rates of invasive tumors. Inflammatory cells can be a factor in carcinogenesis.

Leptin-specific patterns of gene expression have been studied in white adipose tissue. Leptin is a hormone that regulates body weight by various means. Oligonucleotide microarrays were used to study the response of leptin and of food restriction in wild-type mice, ob/ob mice, and transgenic mice that express a constitutively low level of leptin. An analytical method using k-means clustering was developed and with it genes with similar expression patterns could be grouped. Mice called 'ob/ob mice' are those with mutations in leptin.

The progesterone-Wnt connection is of interest relative to side-branching in the mammary gland, which is a derivative of the ectoderm. The existence of transgenic and gene knockout mice has given a way to study how different hormones regulate cell growth, differentiation and death in mammary gland cells. The signaling pathways can be studied and such study may contribute to understanding of breast cancer.

Huntington's disease (HD) and reversal of neuropathology and motor dysfunction in a conditional model of the disease are discussed. Neurodegenerative disorders of this type are characterized by progressive and what is thought to be irreversible clinical and neuropathological symptoms. A tetracycline-regulatable system with mice expressing a mutated huntingtin fragment has been used to set up the conditional HD model. The mice show neuronal inclusions, characteristic neuropathology and progressive motor dysfunction. Blockage of expression in mice with symptoms leads to disappearance of inclusions and amelioration of the behavioral phenotype. Continuous inflow of a mutant protein is needed to maintain inclusions and symptoms. There is thus some possibility that HD could be reversed.

The reversal of neurodegeneration and the work of Rene Hen and colleagues are discussed in this review article. Topics include glutamine repeat diseases and progressive neurodegeneration, and whether impaired protein clearance causes neurodegeneration. A model of reversible neurodegeneration using a line of mice with a transgene with the tetO bidirectional promoter with a lacZ reporter gene on one side and exon1 of huntingtin having 94 CAG repeats (HD94) on the other has been studied. Therapeutic potential is considered.

INK4a/ARF mutations have been found to speed lymphomagenesis and promote chemoresistance by disabling p53. The impact of INK4a/ARF mutations on tumor development and therapy have been studied through use of E-mu-myc transgenic mice. The mice constituitively express c-Myc in the B-cell liage and develop B-cell lymphoma with associated leukemia. There are several reasons for using the mouse model. Human systems are much more difficult to work with.

Transgenic mice with Alzheimer presenilin 1 (PS-1) mutations have been shown to have accelerated neurodegeneration without amyloid plaque formation so the pathogenic role of the PS-1 mutation is upstream of the amyloid cascade. Careful quantitative studies on PS-1 transgenic mice were carried out. Without amyloid plaque formation, neurodegeneration was seen to be accelerated significantly in mice more than 13 months old and having familial Alzheimer disease mutant PS-1.

Transcriptional oscillation of Lunatic fringe (Lfng) has been found to be necessary for somitogenesis. To investigate functional significance of oscillating Lfng activity in somite patterning, transgenic mice were generated. They expressed Lfng in a nonoscillating way in t he presomitic mesoderm using a part of the Delta1 promoter (msf) directing h eterologous gene expression into the paraxial mesoderm.

The transcription factor CCAAT enhancer binding protein [alpha] (C/ EBP [alpha]) is expressed at high levels in liver and adipose tissue. Cell culture studies show that C/EBP [alpha] is sufficient to trigger differentiation of preadipocytes into mature adipocytes, suggesting a central role for C/EBP [alpha] in the development of adipose tissue. C/EBP [alpha] knockout mice die within 7-12 h after birth. Defective gluconeogenesis of the liver and subsequent hypoglycemia contribute to the early death of these animals. This short life span impairs investigation of the development of adipose tissue in these mice. To improve the survival of C/EBP [alpha]-/- animals, we generated a transgenic line that expresses C/EBP [alpha] under the control of the albumin enhancer/promoter. This line was bred into the knockout strain to generate animals that express C/EBP [alpha] in the liver but in no other tissue. The presence of the transgene improved survival of C/EBP [alpha]-/- animals almost 3-fold. Transgenic C/EBP [alpha]-/- animals at 7 days of age show an absence of s.c., perirenal, and epididymal white fat despite excess lipid substrate in the serum, whereas brown adipose tissue is somewhat hypertrophied and shows minimal biochemical alterations. Interestingly, mammary gland fat tissue is present and exhibits normal morphology. The absence of white adipose tissue in many depots in the presence of high serum lipid levels shows that C/EBP [alpha] is required for the in vivo development of this tissue. In contrast, brown adipose tissue differentiation is independent of C/EBP [alpha] expression. The presence of lipid in brown adipose tissue serves as an internal nutritional control, indicating that neither nutritional intake nor lipoprotein composition is likely responsible for the absence of white fat.