Impaired water maze learning performance without altered dopaminergic function in mice heterozygous for the GDNF mutation.

Exogenous glial cell line-derived neurotrophic factor (GDNF) exhibits potent survival-promoting effects on dopaminergic neurons of the nigrostriatal pathway that is implicated in Parkinson's disease and also protects neurons in forebrain ischemia of animal models. However, a role for endogenous GDNF in brain function has not been established. Although mice homozygous for a targeted deletion of the GDNF gene have been generated, these mice die within hours of birth because of deficits in kidney morphogenesis, and, thus, the effect of the absence of GDNF on brain function could not be studied. Herein, we sought to determine whether adult mice, heterozygous for a GDNF mutation on two different genetic backgrounds, demonstrate alterations in the nigrostriatal dopaminergic system or in cognitive function. While both neurochemical and behavioural measures suggested that reduction of GDNF gene expression in the mutant mice does not alter the nigrostriatal dopaminergic system, it led to a significant and selective impairment of performance in the spatial version of the Morris water maze. A standard panel of blood chemistry tests and basic pathological analyses did not reveal alterations in the mutants that could account for the observed performance deficit. These results suggest that endogenous GDNF may not be critical for the development and functioning of the nigrostriatal dopaminergic system but it plays an important role in cognitive abilities.

The skeletal structure of insulin-like growth factor I-deficient mice.

The importance of insulin-like growth factor I (IGF-I) for growth is well established. However, the lack of IGF-I on the skeleton has not been examined thoroughly. Therefore, we analyzed the structural properties of bone from mice rendered IGF-I deficient by homologous recombination (knockout [k/o]) using histomorphometry, peripheral quantitative computerized tomography (pQCT), and microcomputerized tomography (muCT). The k/o mice were 24% the size of their wild-type littermates at the time of study (4 months). The k/o tibias were 28% and L1 vertebrae were 26% the size of wild-type bones. Bone formation rates (BFR) of k/o tibias were 27% that of the wild-type littermates. The k/o bones responded normally to growth hormone (GH; 1.7-fold increase) and supranormally to IGF-I (5.2-fold increase) with respect to BFR. Cortical thickness of the proximal tibia was reduced 17% in the k/o mouse. However, trabecular bone volume (bone volume/total volume [BV/TV]) was increased 23% (male mice) and 88% (female mice) in the k/o mice compared with wild-type controls as a result of increased connectivity, increased number, and decreased spacing of the trabeculae. These changes were either less or not found in L1. Thus, lack of IGF-I leads to the development of a bone structure, which, although smaller, appears more compact.

Hypertension and endothelial dysfunction in apolipoprotein E knockout mice.

Mice lacking ApoE (Apoe(-/-)) develop initially hypercholesterolemia and lastly atherosclerosis. This study examined hemodynamics and endothelial function in 6-week-old Apoe(-/-) mice with hypercholesterolemia only, 7.5-months-old Apoe(-/-) mice with both hypercholesterolemia and atherosclerosis, and age matched controls. One day after implantation of catheters into the carotid artery, arterial pressure was measured in conscious, unrestrained mice. Compared with the respective controls, there was a significant increase in arterial pressure and the ratio of left ventricular weight to body weight in 7.5-month-old Apoe(-/-) mice but not in 6-week-old Apoe(-/-) mice. Histopathological analysis demonstrated significant renal artery disease in the form of extensive atheromatous plaques only in 7.5-month-old Apoe(-/-) mice, whereas no atherosclerotic lesions were found in 6-week-old Apoe(-/-) mice. For evaluation of endothelial function, a laser Doppler perfusion imager with a computer-controlled optical scanner was used to measure cutaneous blood perfusion on the dorsal side of one hind paw before and after topical application of mustard oil, which is known to induce nitric oxide-mediated vasodilation. The mustard oil treatment elicited a substantial increase in blood perfusion (P<0.01), which was similar between 6-week-old Apoe(-/-) mice and controls but significantly blunted in 7.5-month-old Apoe(-/-) mice versus control mice, suggesting nitric oxide-mediated vasodilation is diminished in 7.5-month-old Apoe(-/-) mice but not in 6-week-old Apoe(-/-) mice. In contrast, the increase in blood perfusion induced by topical administration of cilostazol, which induces vasodilation via cyclic adenosine monophosphate, was not different between 7.5-month-old Apoe(-/-) mice and controls. Thus hypertension and endothelial dysfunction observed in 7.5-month-old Apoe(-/-) mice may be due mainly to atherosclerosis.

Somatotroph and lactotroph changes in the adenohypophyses of mice with disrupted insulin-like growth factor I gene.

Pituitary is influenced by circulating and locally produced insulin-like growth factor I (IGF-I). To further elucidate the role of pituitary IGF-I, we compared pituitary morphology of homozygous (IgfI-/-), heterozygous (IgfI+/+), and wild-type (IgfI+/+) fetal and adult mice using light microscopy, immunocytochemistry, in situ hybridization and electron microscopy. In pituitaries of Igf1-/- and Igf1+/- fetal mice (day 18.5) GH RNA signal was decreased. In Igf1-/- adult females, GH cells were significantly diminished in size; GH RNA signal was stronger in Igf1-/- mice compared with IgfI+/+ mice, and the somatotrophs had ultrastructural features of stimulation. The number of PRL cells and PRL hybridization signal were significantly decreased, however plasma PRL levels were elevated in both genders. No changes in other cell types in Igf1-/- mice, and no alterations in Igf1+/- mice were evident. IGF-I treatment for 2 weeks of Igf1-/- mice increased significantly body weights, decreased GH hybridization signal, and had no effect on PRL cells, or PRL plasma levels, whereas in IgfI+/+ mice, PRL RNA signal and PRL plasma levels were markedly increased. In conclusion, IGF-I plays no role in differentiation of pituitary cells, affects the size of somatotrophs in females, and is a stimulator of lactotrophs in both genders.

Effects of Igf1 gene deletion on postnatal growth patterns.

This study documents the temporal and organ-specific effects of Igf1 gene deletion on postnatal growth patterns. Igf1-/- mice are 63+/-4% the size of wildtype (wt) littermates at birth and this ratio persists through postnatal day 20 (P20). After P20, Igf1-/- mice virtually stop growing, while wt littermates double in size from P20 to P40, after which their growth markedly decelerates. As a result, 'full-grown' Igf1-/- mice are less than one third the size of wt littermates. Igf1 gene deletion has disproportionate effects on organ growth. For example, at P10 and P40, Igf1-/- body weights are 63% and 31% of wt, respectively, while Igf1-/- lungs weigh only 34% and 22% of wt at these ages. In contrast, the Igf1-/- heart is disproportionately enlarged, representing approximately 85% of wt at P10 and approximately 56% at P40. Igf1-/- kidney, spleen and liver are slightly but significantly increased in size relative to the degree of reduction in Igf1-/- body weight. These data demonstrate that Igf1 has two major phases or modes of growth promotion. There is an early, growth hormone (GH)-independent Igf1 growth augmentation during perinatal development, responsible for about 35% of growth prior to P20. Then there are later effects due to GH-induced Igf1, which are responsible for increasing animal size by approximately 100% between P20 and 40. The fact that there is virtually no GH-induced growth in the Igf1-/- mice supports the view that Igf1 mediates GH's major effects on somatic growth. Finally, this study shows that Igf1 has discordant effects on pulmonary and cardiac growth parameters, with relative hypoplasia of Igf1-/- lungs and hypertrophy of Igf1-/- hearts.

Insulin-like growth factor I regulates renal development in rodents.

Blocking the action of insulin-like growth factor I (IGF I) impairs kidney development in vitro. However, no renal developmental abnormalities have been reported in newborn transgenic mice that do not express IGF I (Igf1-/-) mice. Ninety-five percent of Igf1-/- mice die immediately following birth. Kidney development continues following birth in rodents. To readdress the question of the participation of IGF I in the process of kidney development, we measured nephron numbers in developed kidneys from Igf1-/- mice that survived past birth, and using a second model of kidney development, characterized the effect of IGF I infused into rat hosts on the renal function of transplanted metanephroi. Igf1-/- mice were born with grossly normal kidneys. At 77 +/- 10 days after birth, Igf1-/- mice that survived were approximately 28% the weight of wild-type (WT) littermates and had proportionally smaller kidneys. The number of nephrons per kidney was reduced by approximately 20% in Igf1-/- mice. Glomerular size was also reduced in Igf1-/- mice. In untreated host rats, neither the size nor inulin clearance of transplanted metanephroi changed significantly from 12-28 weeks postimplantation. The administration of IGF I to hosts did not affect the size of transplanted metanephroi measured at 12-16 weeks following implantation. However, inulin clearances were increased significantly by the administration of IGF I to hosts. Our findings 1) indicate that IGF I plays a role in determining nephron number, 2) suggest that it enhances function in developing kidneys, and 3) establish the potential for the pharmacological use of IGF I to enhance the growth and function of transplanted metanephroi.

Evidence that insulin-like growth factor I and growth hormone are required for prostate gland development.

Insulin-like growth factor I (IGF-I) has been implicated as a factor that may predispose one to prostate cancer. However, no specific relationship between IGF-I and prostate development or cancer in vivo has been established. To determine whether IGF-I was important in prostate development, we examined prostate architecture in IGF-I(-/-) null mice and wild-type littermates. Glands from 44-day-old IGF-I-deficient animals were not only smaller than those from wild-type mice, but also had fewer terminal duct tips and branch points and deficits in tertiary and quaternary branching (P < 0.0001), indicating a specific impairment in gland structure. Administration of des(1-3)-IGF-I for 7 days partially reversed the deficit by increasing those parameters of prostate development (P < 0.006). That IGF-I production probably mediates an effect of GH in this process was indicated by the observations that GH antagonist transgenic mice also had significantly impaired prostate development (P < 0.0002) and that bovine GH had no independent effect on stimulating prostate development in IGF-I null animals. The data indicate that IGF-I deficiency is the proximate cause of impaired prostate development and give credence to the idea that, like testosterone, GH and IGF-I may be involved in prostate cancer growth as an extension of a normal process.

IGF-I deficient mice show reduced peripheral nerve conduction velocities and decreased axonal diameters and respond to exogenous IGF-I treatment.

Although insulin-like growth factor-I (IGF-I) can act as a neurotrophic factor for peripheral neurons in vitro and in vivo following injury, the role IGF-I plays during normal development and functioning of the peripheral nervous system is unclear. Here, we report that transgenic mice with reduced levels (two genotypes: heterozygous Igf1+/- or homozygous insertional mutant Igf1m/m) or totally lacking IGF-I (homozygous Igf1-/-) show a decrease in motor and sensory nerve conduction velocities in vivo. In addition, A-fiber responses in isolated peroneal nerves from Igf1+/- and Igf1-/- mice are impaired. The nerve function impairment is most profound in Igf1-/- mice. Histopathology of the peroneal nerves in Igf1-/- mice demonstrates a shift to smaller axonal diameters but maintains the same total number of myelinated fibers as Igf1+/+ mice. Comparisons of myelin thickness with axonal diameter indicate that there is no significant reduction in peripheral nerve myelination in IGF-I-deficient mice. In addition, in Igf1m/m mice with very low serum levels of IGF-I, replacement therapy with exogenous recombinant hIGF-I restores both motor and sensory nerve conduction velocities. These findings demonstrate not only that IGF-I serves an important role in the growth and development of the peripheral nervous system, but also that systemic IGF-I treatment can enhance nerve function in IGF-I-deficient adult mice.

Insulin-like growth factor 1 is required for G2 progression in the estradiol-induced mitotic cycle.

Insulin-like growth factor 1 (IGF1) has been proposed as a "G1-progression factor" and as a mediator of estradiol's (E2) mitogenic effects on the uterus. To test these hypotheses, we compared E2's mitogenic effects on the uteri of Igf1-targeted gene deletion (null) and wild-type littermate mice. The proportion of uterine cells involved in the cell cycle and G1- and S-phase kinetics were not significantly different in wild-type and Igf1-null mice. However, the appearance of E2-induced mitotic figures and cell number increases were profoundly retarded in Igf1-null uterine tissue. There was a significant increase in nuclear DNA concentration in Igf1-null cells, consistent with a G2 arrest. Interestingly, apoptotic cells were also significantly reduced in abundance, and the normal massive apoptotic response to E2 withdrawal was absent in the Igf1-null uterus. These data show that Igf1 is an essential mediator of E2's mitogenic effects, with a critical role not in G1 progression but in G2 progression.

The role of the growth hormone/insulin-like growth factor I axis in stimulation of protein synthesis in skeletal muscles following oral refeeding.

The mechanisms behind stimulation of protein synthesis in skeletal muscles following oral feeding are not well understood. Previous research has not confirmed that insulin is a major factor behind this stimulation. In the present study we have used genetically altered mice, with either a lack of GH secretion due to a mutational gene inactivation [GH (-/-) dwarf, DW/JOrlBom-dw] or mice with a homozygous site-specific insertion mutation in the insulin-like growth factor-1 gene [IGF-I (m/m)], leading to a deficient IGF-I production. These gene knock-outs were used in comparison to their normal wild types for evaluation of the role that the GH/IGF-I axis may have in activation of nutritionally induced stimulation of protein synthesis in skeletal muscles during oral refeeding. Weight stable adult C57B16 mice served as an additional normal control group. Protein synthesis was measured by a modified flooding dose technique with radioactive L-[14C-U]phenylalanine incorporation into acid precipitated muscle proteins. Fractional protein synthesis in skeletal muscles after an overnight fast was comparable among C57B16 (0.076 +/- 0.009%/h), wild-type IGF-I(+/+) (0.061 +/- 0.008) and IGF-I(m/m) deficient mice (0.068 +/- 0.006%/h), whereas GH(-/-) incompetent mice had a lower fractional synthesis rate compared with GH(+/+) competent mice (0.045 +/- 0.006 vs. 0.068 +/- 0.007, P < 0.05). Refeeding with standard chow diet stimulated protein synthesis in muscles by more than 60% in all animal groups. This response was independent of circulating GH, total IGF-I concentrations in blood, as well as up-regulation of locally produced IGF-I messenger RNA (mRNA) in skeletal muscles.

A mouse model of human familial hypercholesterolemia: markedly elevated low density lipoprotein cholesterol levels and severe atherosclerosis on a low-fat chow diet.

Mutations in the low density lipoprotein (LDL) receptor gene cause familial hypercholesterolemia, a human disease characterized by premature atherosclerosis and markedly elevated plasma levels of LDL cholesterol and apolipoprotein (apo) B100. In contrast, mice deficient for the LDL receptor (Ldlr-/-) have only mildly elevated LDL cholesterol levels and little atherosclerosis. This difference results from extensive editing of the hepatic apoB mRNA in the mouse, which limits apoB100 synthesis in favor of apoB48 synthesis. We have generated Ldlr-/- mice that cannot edit the apoB mRNA and therefore synthesize exclusively apoB100. These mice had markedly elevated LDL cholesterol and apoB100 levels and developed extensive atherosclerosis on a chow diet. This authentic model of human familial hypercholesterolemia will provide a new tool for studying atherosclerosis.

Biochemical and morphometric analyses show that myelination in the insulin-like growth factor 1 null brain is proportionate to its neuronal composition.

To elucidate the role of insulin-like growth factor 1 (IGF1) in the normal development of brain myelination, we used behavioral, biochemical, and histological analyses to compare the myelination of brains from Igf1(-/-) and wild-type (WT) littermate mice. The studies were conducted at postnatal day 40, at which time the Igf1(-/-) mice weighed approximately 66% less than wild-type mice. However, the Igf1(-/-) brain weight was only reduced by approximately 34%. Formal neurological testing showed no sign of central or peripheral myelinopathy in Igf1(-/-) mice. Myelin composition was not significantly different, and myelin concentration, normalized to brain weight or protein, was equal in Igf1(-/-) and WT mice. Likewise, concentrations of myelin-specific proteins (MBP, myelin proteolipid protein, MAG, and 2',3'-cyclic nucleotide,3'-phosphodiesterase) were not significantly different in Igf1(-/-) and WT mice. The myelin-associated lipids galactocerebroside and sulfatide were modestly reduced in Igf1(-/-) brains. Regional oligodendrocyte populations and myelin staining patterns were comparable in Igf1(-/-) and WT brains, with the notable exception of the olfactory system. The Igf1(-/-) olfactory bulb was profoundly reduced in size and was depleted of mitral neurons and oligodendrocytes, and its efferent tracts were depleted of myelin. In summary, this study shows that myelination of the Igf1(-/-) brain is proportionate to its neuronal composition. Where projection neurons are preserved despite the deletion of IGF1, as in the cerebellar system, oligodendrocytes and myelination are indistinguishable from wild type. Where projection neurons are depleted, as in the olfactory bulb, oligodendrocytes are also depleted, and myelination is reduced in proportion to the reduced projection neuron mass. These data make a strong case for the primacy of axonal factors, not including IGF1, in determining oligodendrocyte survival and myelination.

Primary B cell development is impaired in mice with defects of the pituitary/thyroid axis.

There has been considerable speculation that hormones produced in the anterior pituitary gland act as positive regulators of primary B cell development in the bone marrow. In order to identify endocrine factors that have such a role, B cell differentiation was examined in a panel of mice with genetic mutations that result in compromised production of one or more anterior pituitary hormones. This analysis demonstrated that the frequency of B lineage cells is significantly reduced in the dwarf and hypothyroid strains of mice, which have defects in the pituitary/thyroid axis, and that the production of normal numbers of pre-B cells is particularly dependent upon thyroid hormones. B cell development was normal in Little and IGF-I knockout animals, which have defects in the production of growth hormone and/or insulin-like growth factor I. The dependence of B lymphopoiesis on thyroid hormones appeared to be specific for that lineage, as myelopoiesis and thymopoiesis were normal in dwarf and hypothyroid mice. In addition to describing a specific endocrine hormone involved in the regulation of B cell development, these data provide evidence that normal production of bone marrow B lineage cells is dependent on extramedullary signals.

Peripheral vascular stenosis in apolipoprotein E-deficient mice. Potential roles of lipid deposition, medial atrophy, and adventitial inflammation.

A systematic analysis of the distribution of advanced atherosclerotic lesions was undertaken in chow-fed, 9-month-old apolipoprotein (apo) E-deficient mice to identify sites amenable for study of mechanisms of formation of stenotic lesions. The arterial tree was dissected intact and included medium-sized arteries in the extremities as well as arteries of the head and neck. The most reproducible lesions were seen in the ascending aorta and in the carotid, femoral, and popliteal arteries. Casting of the vascular tree provided additional verification of the presence of lumen narrowing in the external branches of the carotid artery. Consistent with what has been observed in human atherosclerotic arteries, there was dilation in response to lesion growth and no correlation between lesion mass and lumen loss in the mouse arteries. This adaptation was especially true in the ascending aorta, where normal lumen size was maintained at atherosclerotic sites. In contrast, the external carotid arteries were stenotic in 9 of 12 animals. Here too, however, loss of lumen did not correlate with lesion mass but did correlate with adventitial inflammation and medial atrophy. Lumen narrowing also occurred most frequently at sites where extracellular cholesterol clefts were a prominent part of the lesion. These data suggest that the stenotic process in advanced atherosclerotic vessels may depend on death of medial smooth muscle cells, possibly in response to inflammatory changes in the plaque or adventitia.

Elevated blood pressure and enhanced myocardial contractility in mice with severe IGF-1 deficiency.

To circumvent the embryonic lethality of a complete deficiency in insulin-like growth factor 1 (IGF-1), we generated mice homozygous for a site-specific insertional event that created a mutant IGF-1 allele (igf1m). These mice have IGF-1 levels 30% of wild type yet survive to adulthood, thereby allowing physiological analysis of the phenotype. Miniaturized catheterization technology revealed elevated conscious blood pressure in IGF-1(m/m) mice, and measurements of left ventricular contractility were increased. Adenylyl cyclase activity was enhanced in IGF-1(m/m) hearts, without an increase in beta-adrenergic receptor density, suggesting that crosstalk between IGF-1 and beta-adrenergic signaling pathways may mediate the increased contractility. The hypertrophic response of the left ventricular myocardium in response to aortic constriction, however, was preserved in IGF-1(m/m) mice. We conclude that chronic alterations in IGF-1 levels can selectively modulate blood pressure and left ventricular function, while not affecting adaptive myocardial hypertrophy in vivo.

Heterozygous embryonic lethality induced by targeted inactivation of the VEGF gene.

Angiogenesis is required for a wide variety of physiological and pathological processes. The endothelial cell-specific mitogen vascular endothelial growth factor (VEGF) is a major mediator of pathological angiogenesis. Also, the expression of VEGF and its two receptors, Flt-1 and Flk-1/KDR, is related to the formation of blood vessels in mouse and rat embryos. Mice homozygous for mutations that inactivate either receptor die in utero between days 8.5 and 9.5. However, ligand(s) other than VEGF might activate such receptors. To assess the role of VEGF directly, we disrupted the VEGF gene in embryonic stem cells. Here we report the unexpected finding that loss of a single VEGF allele is lethal in the mouse embryo between days 11 and 12. Angiogenesis and blood-island formation were impaired, resulting in several developmental anomalies. Furthermore, VEGF-null embryonic stem cells exhibit a dramatically reduced ability to form tumours in nude mice.

Targeted disruption of the mouse apobec-1 gene abolishes apolipoprotein B mRNA editing and eliminates apolipoprotein B48.

A site-specific C to U editing reaction modifies nuclear apolipoprotein B100 (apoB100) mRNA, producing apolipoprotein B48 in the mammalian small intestine. This reaction is mediated by a multicomponent enzyme complex, which contains a catalytic subunit, Apobec-1. We have used gene targeting to disrupt mouse apobec-1 in order to establish its requisite importance in apoB mRNA editing and also, in view of its widespread tissue distribution in rodents, as a preliminary indication of other potential roles. Both heterozygous (apobec-1+/-) and homozygous (apobec-1-/-) gene-targeted mice appear healthy and fertile with no alterations in serum cholesterol or triglyceride concentrations. The apobec-1+/- mice demonstrated reduced levels of hepatic apoB mRNA editing. By contrast, levels of small intestinal apoB mRNA editing were indistinguishable in wild-type and apobec-1+/- animals, suggesting that Apobec-1 is expressed in limited quantities in the liver but not in the small intestine. The apobec-1-/- mice lacked detectable levels of Apobec-1 mRNA, expressed only unedited apoB mRNA in all tissues, and contained no apoB48 in their serum, demonstrating that there is no functional duplication of this gene.

Igf1 gene disruption results in reduced brain size, CNS hypomyelination, and loss of hippocampal granule and striatal parvalbumin-containing neurons.

Homozygous Igf1-/- mice at 2 months of age had reduced brain weights, with reductions evenly affecting all major brain areas. The gross morphology of the CNS was normal, but the size of white matter structures in brain and spinal cord was strongly reduced, owing to decreased numbers of axons and oligodendrocytes. Myelinated axons were more strongly reduced in number than unmyelinated axons. The volume of the dentate gyrus granule cell layer was reduced in excess of the decrease in brain weight. Among populations of calcium-binding protein-containing neurons, there was a selective reduction in the number of striatal parvalbumin-containing cells. Numbers of mesencephalic dopaminergic neurons, striatal and basal forebrain cholinergic neurons, and spinal cord motoneurons were unaffected. Cerebellar morphology was unaltered. Our findings suggest cell type- and region-specific functions for IGF-I and emphasize prominent roles in axon growth and maturation in CNS myelination.

Mouse models of IGF-I deficiency generated by gene targeting.

Insulin-like growth factor-I (IGF-I) is a pleiotropic hormone synthesized in a wide variety of cell types. As its name suggests, it is highly homologous to insulin and has many similar growth promoting effects. IGF-I is believed to be the major anabolic mediator of growth hormone action. A second insulin-like growth factor (IGF-II) is expressed at high levels in the fetus. IGF-II is homologous to IGF-I and interacts with the IGF-I (type 1) receptor as well as its own (type 2) receptor. Targeted manipulation of the mouse genome enables dissection of gene function in the context of the whole animal. We have generated strains of mice deficient in IGF-I production by homologous recombination that demonstrate important roles for IGF-I in both pre- and postnatal development.

IGF-I is required for normal embryonic growth in mice.

IGF-I is a pleiotropic hormone reported to affect linear growth, glucose metabolism, organ homeostasis, and the immune and neurologic systems. In contrast to IGF-II, IGF-I is expressed at low levels embryonically and has been thought to be more important for postnatal growth and development. To investigate the role of IGF-I in normal development we generated mice with an inactive IGF-I gene by homologous recombination in ES cells. Heterozygous mice are healthy and fertile, but they are 10-20% smaller than wild-type littermates and have lower than normal levels of IGF-I. The size reduction is attributable to a decrease in organs and muscle and bone mass. However, all tissues appear histologically normal. At birth homozygous mutant mice (IGF-I-/-) are < 60% body weight of wild type. Greater than 95% of IGF-I-/- pups die perinatally. Histopathology is characterized by underdevelopment of muscle tissue. Lungs of late embryonic and neonates also appeared less organized with ill-defined alveolae. IGF-I appears to be essential for correct embryonic development in mice.