Anselm P. D'Costa

Pigment epithelium-derived factor promotes the survival and differentiation of developing spinal motor neurons.

Pigment epithelium‐derived factor (PEDF) is a member of the serine protease inhibitor (serpin) superfamily that has been shown previously to promote the survival and/or differentiation of rat cerebellar granule neurons and human retinoblastoma cells in vitro. However, in contrast to most serpins, PEDF has no inhibitory activity against any known proteases, and its described biological activities do not appear to require the serpin‐reactive loop located toward the carboxy end of the polypeptide. Because another serpin, protease nexin‐1, has been shown to promote the in vivo survival and growth of motor neurons, the authors investigated the potential neurotrophic effects of PEDF on spinal cord motor neurons in highly enriched cultures and in vivo after injury. Here, it is shown that native bovine and recombinant human PEDF promoted the survival and differentiation (neurite outgrowth) of embryonic chick spinal cord motor neurons in vitro in a dose‐dependent manner. A truncated form of PEDF that lacks ≈62% of the carboxy end of the polypeptide comprising the homologous serpin‐reactive loop also exhibited neurotrophic activities similar to those of the full‐length protein. Furthermore, the data here showed that PEDF was transported retrogradely and prevented the death and atrophy of spinal motor neurons in the developing neonatal mouse after axotomy. These results indicate that PEDF exerts trophic effects on motor neurons, and, together with previous reports, these findings suggest that this protein may be useful as a pharmacologic agent to promote the development and maintenance of motor neurons. J. Comp. Neurol. 412:506–514, 1999. Published 1999 Wiley‐Liss, Inc.

Expression of insulin-like growth factor-1 (IGF-1) and IGF-binding protein 2 (IGF-BP2) in the hippocampus following cytotoxic lesion of the dentate gyrus.

Receptor binding and gene expression of several members of the IGF gene family were examined in the rat brain following lesion of the hippocampal dentate gyrus granular cells by intradentate colchicine injection. Dentate granular cell loss was accompanied by extensive reactive gliosis in the lesioned hippocampus and damaged overlying cortex, as verified by the increase in GFAP mRNA and BS‐1 lectin binding. At 4 days post‐lesion, 125I‐IGF‐2 binding was dramatically increased within the lesioned dentate gyrus and damaged overlying cortex, and corresponded temporally and anatomically with increased IGF‐BP2 gene expression following the lesion. Increased IGF‐BP3 gene expression was only observed in the overlying cortex at 10 days post‐lesion, and corresponded with an increase in 125I‐IGF‐1 binding at the injured surface of the cortex. Type‐2 IGF receptor mRNA expression was reduced to background levels in the lesioned dentate gyrus, suggesting that IGF‐BP2 was a major component of the observed increase in 125I‐IGF‐2 binding. In situ hybridization also revealed a prominent increase in IGF‐1 mRNA expression by 4 days post‐lesion, which was localized within the lesioned dentate gyrus and damaged cortical areas, and was shown to be expressed by microglia. While no IGF‐2 mRNA expression was observed within the CNS, either prior to, or following the lesion, IGF‐2 mRNA expression was observed in the choroid plexus, meningeal membranes, and in blood vessel endothelium, providing a potential source for the transport of IGF‐2 into the CNS. In the injured CNS, increased IGF‐BP2 expression may act to maintain or transport IGF‐1 or IGF‐2, as well as modulate the local autocrine and paracrine actions of the IGFs. Increased microglial IGF‐1 expression following colchicine treatment correlates with the timing of a number of post‐traumatic events within the CNS, suggesting that IGF‐1 may have a role as a neuroprotectant for surviving neurons and signal for local neuronal sprouting, as well as a role in reactive astrogliosis.

Moderate caloric restriction increases type 1 IGF receptors and protein synthesis in aging rats

Insulin-like growth factor-1 (IGF-1) is an anabolic hormone that mediates the actions of growth hormone (GH) and is found at lower concentrations in aged animals. These decreases in GH and IGF-1 appear to have important physiological consequences for aging, since protein synthesis decreases with age, and administration of GH and/or IGF-1 has been shown to increase protein synthesis. The present study was designed to determine (a) the relationship between the age-related changes in rates of tissue protein synthesis and plasma IGF-1 concentrations, (b) type 1 IGF receptor density in tissues and (c) whether long-term moderate caloric restriction, which is known to increase life-span, affects these relationships. Male Brown Norway rats were fed ad libitum or caloric-restricted (60% ad libitum) from 14 weeks of age and sacrificed at different ages. In ad libitum fed animals there were age-related decreases in plasma IGF-1 concentrations (14%) and in the rates of protein synthesis of the heart (36%) and liver (38%). Type 1 IGF receptor density remained constant in all tissues with age. The caloric-restricted animals exhibited plasma IGF-1 concentrations 33 to 42% lower than the ad libitum fed animals. However, rates of protein synthesis increased by 70 and 30% in heart and diaphragm, and this increase was associated with 60 to 100% increases in type 1 IGF receptor densities when compared with ad libitum fed animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanisms of insulin-like growth factor regulation of programmed cell death of developing avian motoneurons

During development of the avian neuromuscular system, lumbar spinal motoneurons (MNs) innervate their muscle targets in the hindlimb coincident with the onset and progression of MN programmed cell death (PCD). Paralysis (activity blockade) of embryos during this period rescues large numbers of MNs from PCD. Because activity blockade also results in enhanced axonal branching and increased numbers of neuromuscular synapses, it has been postulated that following activity blockade, increased numbers of MNs can gain access to muscle-derived trophic agents that prevent PCD. An assumption of the access hypothesis of MN PCD is the presence of an activity-dependent, muscle-derived sprouting or branching agent. Several previous studies of sprouting in the rodent neuromuscular system indicate that insulin-like growth factors (IGFs) are candidates for such a sprouting factor. Accordingly, in the present study we have begun to test whether the IGFs may play a similar role in the developing avian neuromuscular system. Evidence in support of this idea includes the following: (a) IGFs promote MN survival in vivo but not in vitro; (b) neutralizing antibodies against IGFs reduce MN survival in vivo; (c) both in vitro and in vivo, IGFs increase neurite growth, branching, and synapse formation; (d) activity blockade increases the expression of IGF-1 and IGF-2 mRNA in skeletal muscles in vivo; (e) in vivo treatment of paralyzed embryos with IGF binding proteins (IGF-BPs) that interfere with the actions of endogenous IGFs reduce MN survival, axon branching, and synapse formation; (f) treatment of control embryos in vivo with IGF-BPs also reduces synapse formation; and (g) treatment with IGF-1 prior to the major period of cell death (i.e., on embryonic day 6) increases subsequent synapse formation and MN survival and potentiates the survival-promoting actions of brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) administered during the subsequent 4- to 5-day period of PCD. Collectively, these data provide new evidence consistent with the role of the IGFs as activity-dependent, muscle-derived agents that play a role in regulating MN survival in the avian embryo.

Insulin-like growth factor-1 stimulation of protein synthesis is attenuated in cerebral cortex of aging rats

It has been postulated that brain aging and the accompanying neurodegenerative processes associated with aging result from a deterioration of mechanisms that regulate the maintenance of basic cellular processes. In the present study, it was hypothesized that decreased availability and/or diminished responsiveness of tissues to growth factors such as insulin-like growth factor-1 may be partly responsible for decreases in total protein synthesis previously observed in aging animals. Male Brown Norway rats (5-7 and 27-28 months old) were used to determine (1) whether in vivo protein synthesis in cortex, hippocampus, hypothalamus and cerebellum decreases with age and (2) whether these deficiencies are associated with age-related alterations in response to insulin-like growth factor-1, des (1-3) IGF-1 or insulin. Analysis of in vivo protein synthesis rates revealed a decline of 20% in cortex of old rats (P < 0.05) but no changes were observed in hippocampus, hypothalamus, or cerebellum. Stimulation of cortical slices in vitro with insulin-like growth factor-1, des (1-3) insulin-like growth factor-1, or insulin increased protein synthesis rates in young animals, but the response to these growth factors was blunted in old animals. Analysis of type 1 insulin-like growth factor receptor densities by quantitative autoradiography demonstrated age-related decreases in receptor levels in cerebellar cortex and dentate gyrus of the hippocampus but no changes in cortex. Regional distribution of type 1 insulin-like growth factor receptors within each of these tissues did not appear to change with age.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of in utero ethanol exposure on the postnatal ontogeny of insulin-like growth factor-1, and type-1 and type-2 insulin-like growth factor receptors in the rat brain

There is convincing evidence that alcohol consumption during pregnancy causes major CNS abnormalities; however, the molecular and cellular basis of these dysfunctions is currently not understood. This study examined the effects of prenatal ethanol exposure on the expression of insulin-like growth factor-1 messenger RNA and type-1 and type-2 receptor protein and messenger RNA expression in the developing rat brain. Mothers were maintained on an ethanol containing liquid diet from day 2 of pregnancy through parturition and the offspring were killed at birth, 10, 20 and 40 days of age. Insulin-like growth factor-1 messenger RNA, and insulin-like growth factor receptors demonstrated developmentally dependent expression in specific brain regions throughout the postnatal period of CNS maturation. Insulin-like growth factor-1 gene expression in the brain, as analysed by dot-blot hybridization, was greatest at birth, and decreased 61% in ad libitum and pair-fed animals by 20 days of age. In contrast, ethanol-treated animals exhibited only a 25% decrease in insulin-like growth factor-1 messenger RNA levels during the same period. This delay in insulin-like growth factor-1 messenger RNA maturation may be related to a developmental delay in CNS development in the prenatally ethanol exposed offspring. Prenatal ethanol exposure did not alter the observed localization of insulin-like growth factor-1 messenger RNA. While alterations were observed in long-term insulin-like growth factor-1 messenger RNA regulation, quantitative receptor autoradiography and in situ hybridization demonstrated no alterations in either type-1 or type-2 insulin-like growth factor receptor populations in ethanol-treated animals. Changes in hepatic and plasma insulin-like growth factor-1 and insulin-like growth factor-binding protein regulation have also been observed in these animals, suggesting changes in protein translation and the autocrine/paracrine actions of this peptide. The present study demonstrated that insulin-like growth factor-1 messenger RNA and insulin-like growth factor receptors are regionally expressed during early postnatal development and that ethanol administration influenced the long-term regulation of insulin-like growth factor messenger RNA levels in the brain without affecting either its localization or insulin-like growth factor receptor populations.(ABSTRACT TRUNCATED AT 400 WORDS)

Distribution of Insulin-Like Growth Factor 1 (IGF-1) and 2 (IGF-2) Receptors in the Hippocampal Formation of Rats and Mice

This study demonstrated species differences in IGF-1 and IGF-2 receptor binding and localization in the hippocampus of the rat and mouse. Competition binding studies indicated that there were no differences in the relative binding affinities for the type 1 or type 2 receptors between the brains of these animals. These results suggested that the observed species differences were not attributable to alterations in IGF receptor kinetics. Receptor autoradiographic analyses demonstrated that IGF-1 binding differed in both the localization and overall receptor densities observed, with the rat demonstrating more specific localization and greater receptor density in the hippocampus than the mouse. The rat also exhibited a greater density of IGF-2 receptors in the hippocampus than the mouse. Despite differences in IGF receptor populations, both species exhibit similar hippocampal structure and lamination. Therefore, these results demonstrate a disparity in the localization of IGF receptor binding in the rat and mouse, suggesting that IGFs in these species are differentially regulated, with distinct neuromodulatory, neurotrophic, and/or developmental roles in this region of the brain. Previous comparative anatomical studies of the hippocampal formation of rats and mice fail to offer an explanation for the absence or reduction of binding of IGF-1 in the mouse. Although the mouse has a greater cell density in the s. granulosum than the rat, and both species exhibit similar glia and synaptic contact densities in the s. moleculare of the dentate gyrus, the mouse exhibits a complete absence of IGF-1 binding in this region. The lack of anatomical differences in the hippocampal formation of these species suggests that the patterns observed in IGF binding result from alterations in either neurochemical modulation of these neurons or specific neurotrophic requirements of the cells in this region. Differences have been reported on the concentrations and binding of various neurotransmitters in the hippocampus of these species, however these differences do not easily account for the variations observed in IGF binding in this study. IGFs are known to influence acetylcholine neurotransmission in the hippocampus as well as other brain areas in the rat. Recently, a truncated form of IGF-1, in which a tripeptide is cleaved from the N-terminus of the peptide, has been reported in brain. The cleaved tripeptide has been shown to activate glutamate receptors, which may dramatically influence excitatory neurotransmission in this region. Therefore, in addition to the possible neurotrophic actions of the peptide itself, subsequent processing of IGF-1 may be an important aspect of IGF-1 activity in the brain.(ABSTRACT TRUNCATED AT 400 WORDS)