Regulation Of Neuronal Development Research Articles

Difference of fibroblast growth factor receptor 1 expression among CA1-3 regions of the gerbil hippocampus after transient cerebral ischemia

Fibroblast growth factors are important regulators of neuronal development. In this study, we observed fibroblast growth factor receptor 1 (FGFR1) immunoreactivity and its protein levels in the hippocampus proper (CA1-3 regions) of the gerbil at various time points after ischemia/reperfusion. In the sham-operated group, FGFR1 immunoreaction was not detected in the hippocampus proper. FGFR1 immunoreaction was first detected in non-pyramidal neurons in the CA1-3 region at 12h and 1day after ischemia/reperfusion. From 2days after ischemia/reperfusion, FGFR1 immunoreaction was found in astrocytes, not in microglial cells, in the CA1 region: FGFR1 immunoreactivity and the number of astrocytes were significantly increased at 5days post-ischemia. Western blot analysis revealed that FGFR1 protein levels were also increased from 1day after ischemia/reperfusion. These results indicate that increase of FGFR1 in astrocytes of the ischemic CA1 region may be associated with gliosis followed by delayed neuronal death.

MicroRNAs in the neural system

Precise spatio-temporal control of gene expression at transcriptional and translational levels is required for both of proper developmental programming of the central nervous system and the performing of normal brain functions. Many studies have demonstrated that micro-RNAs (miRNAs), a class of endogenous small RNAs, participate in post-transcriptional regulation of gene expression, and thus execute regulatory functions in various biologic processes. Emerging evidence indicates that miRNAs participate in gene regulatory networks during the developmental, physiologic, and pathological processes of the brain. In this review, we attempt to summarize some of the recent advances in research on the involvement of miRNAs in the regulation of neuronal development, neuroplasticity, and brain diseases, revealing their indispensable roles in neural functions.

Recombinant adeno-associated virus-mediated microRNA delivery into the postnatal mouse brain reveals a role for miR-134 in dendritogenesis in vivo

Recent studies using primary neuronal cultures have revealed important roles of the microRNA pathway in the regulation of neuronal development and morphology. For example, miR-134 is involved in dendritogenesis and spine development in hippocampal neurons by regulating local mRNA translation in dendrites. The in vivo roles of microRNAs in these processes are still uninvestigated, partly due to the lack of tools enabling stable in vivo delivery of microRNAs or microRNA inhibitors into neurons of the mammalian brain. Here we describe the construction and validation of a vector-based tool for stable delivery of microRNAs in vivo by use of recombinant adeno-associated virus (rAAV). rAAV-mediated overexpression of miR-134 in neurons of the postnatal mouse brain provided evidence for a negative role of miR-134 in dendritic arborization of cortical layer V pyramidal neurons in vivo, thereby confirming previous findings obtained with cultured neurons. Our system provides researchers with a unique tool to study the role of any candidate microRNA in vivo and can easily be adapted to microRNA loss-of-function studies. This platform should therefore greatly facilitate investigations on the role of microRNAs in synapse development, plasticity and behavior in vivo.

2-Bromopalmitate and 2-(2-hydroxy-5-nitro-benzylidene)-benzo[b]thiophen-3-one inhibit DHHC-mediated palmitoylation in vitro

Pharmacologic approaches to studying palmitoylation are limited by the lack of specific inhibitors. Recently, screens have revealed five chemical classes of small molecules that inhibit cellular processes associated with palmitoylation (Ducker, C. E., L. K. Griffel, R. A. Smith, S. N. Keller, Y. Zhuang, Z. Xia, J. D. Diller, and C. D. Smith. 2006. Discovery and characterization of inhibitors of human palmitoyl acyltransferases. Mol. Cancer Ther. 5: 1647-1659). Compounds that selectively inhibited palmitoylation of N-myristoylated vs. farnesylated peptides were identified in assays of palmitoyltransferase activity using cell membranes. Palmitoylation is catalyzed by a family of enzymes that share a conserved DHHC (Asp-His-His-Cys) cysteine-rich domain. In this study, we evaluated the ability of these inhibitors to reduce DHHC-mediated palmitoylation using purified enzymes and protein substrates. Human DHHC2 and yeast Pfa3 were assayed with their respective N-myristoylated substrates, Lck and Vac8. Human DHHC9/GCP16 and yeast Erf2/Erf4 were tested using farnesylated Ras proteins. Surprisingly, all four enzymes showed a similar profile of inhibition. Only one of the novel compounds, 2-(2-hydroxy-5-nitro-benzylidene)-benzo[b]thiophen-3-one [Compound V (CV)], and 2-bromopalmitate (2BP) inhibited the palmitoyltransferase activity of all DHHC proteins tested. Hence, the reported potency and selectivity of these compounds were not recapitulated with purified enzymes and their cognate lipidated substrates. Further characterization revealed both compounds blocked DHHC enzyme autoacylation and displayed slow, time-dependent inhibition but differed with respect to reversibility. Inhibition of palmitoyltransferase activity by CV was reversible, whereas 2BP inhibition was irreversible.

Think locally: control of ubiquitin‐dependent protein degradation in neurons

The nervous system coordinates many aspects of body function such as learning, memory, behaviour and locomotion. Therefore, it must develop and maintain an intricate network of differentiated neuronal cells, which communicate efficiently with each other and with non-neuronal target cells. Unlike most somatic cells, differentiated neurons are post-mitotic and characterized by a highly polarized morphology that determines the flow of information. Among other post-translational modifications, the ubiquitination of specific protein substrates was recently shown to have a crucial role in the regulation of neuronal development and differentiation. Here, we review recent findings that illustrate the mechanisms that mediate the temporal and spatial control of neuronal protein turnover by the ubiquitin-proteasome system (UPS), which is crucial for the development and function of the nervous system.

Transcription of the chicken Grin1 gene is regulated by the activity of SP3 and NRSF in undifferentiated cells and neurons

The NMDA (N-methyl-D-aspartate) receptors are important in the regulation of neuronal development, synaptic plasticity, learning and memory, and are involved in several brain pathologies. The NR1 subunit is essential for the assembly of functional receptors, as it forms the calcium-permeable ion channel and contains the obligatory co-agonist binding site. Previous studies have shown that NR1 gene (Grin1) expression is up-regulated during neuronal differentiation and its expression is widespread in the central nervous system. We have previously cloned the chicken Grin1 gene and 1.9 kb of the 5'-regulatory region. In the present study, we analysed the molecular mechanisms that regulate chicken Grin1 gene transcription in undifferentiated cells and neurons. By functional analysis of chicken Grin1-luciferase gene 5'-regulatory region constructs, we demonstrate that the basal promoter is delimited within 210 bp upstream from the main transcription initiation site. DNA-protein binding and functional assays revealed that the 5'-UTR (untranslated region) has one consensus NRSE (neuron-restrictive silencing element) that binds NRSF (neuron-restrictive silencing factor), and one SP (stimulating protein transcription factor) element that binds SP3, both repressing Grin1 gene transcription in undifferentiated P19 cells (embryonic terato-carcinoma cells) and PC12 cells (phaeochromocytoma cells). The promoter region lacks a consensus TATA box, but contains one GSG/SP (GSG-like box near a SP-consensus site) that binds SP3 and up-regulates gene transcription in embryonic chicken cortical neurons. Taken together, these results demonstrate a dual role of SP3 in regulating the expression of the Grin1 gene, by repressing transcription in the 5'-UTR in undifferentiated cells as well as acting as a transcription factor, increasing Grin1 gene transcription in neurons.

Diverse roles of Rho family GTPases in neuronal development, survival, and death

Rho family GTPases (eg., RhoA, Rac1 and Cdc42) are monomeric G-proteins that act as key transducers of extracellular signals to the actin cytoskeleton. In the nervous system, Rho family GTPases are essential regulators of neuronal growth cone motility, axonal migration, and dendritic spine morphogenesis. Given these vital functions, it is perhaps not surprising that mutations in several proteins involved in Rho GTPase signaling are causative in some forms of mental retardation. In addition, numerous recent studies have identified Rho family GTPases as central players in the molecular pathways that determine neuronal survival and death. Interestingly, individual Rho family members have been shown to play either a pro-death or pro-survival role in the nervous system depending on both the type of neuron and the particular neurodegenerative insult involved. This review summarizes current work demonstrating a critical role for Rho family GTPases and their effectors in the regulation of neuronal development, survival, and death. These findings may be particularly relevant in the context of specific neurodegenerative disorders in which Rho family GTPase function is altered, such as loss-of-function of the Rac1 guanine nucleotide exchange factor, alsin, in juvenile-onset amyotrophic lateral sclerosis.

Regulation of Dendritogenesis via a Lipid-Raft-Associated Ca2+/Calmodulin-Dependent Protein Kinase CLICK-III/CaMKIγ

Ca(2+) signaling plays a central role in activity-dependent regulation of dendritic arborization, but key molecular mechanisms downstream of calcium elevation remain poorly understood. Here we show that the C-terminal region of the Ca(2+)/calmodulin-dependent protein kinase CLICK-III (CL3)/CaMKIgamma, a membrane-anchored CaMK, was uniquely modified by two sequential lipidification steps: prenylation followed by a kinase-activity-regulated palmitoylation. These modifications were essential for CL3 membrane anchoring and targeting into detergent-resistant lipid microdomains (or rafts) in the dendrites. We found that CL3 critically contributed to BDNF-stimulated dendritic growth. Raft insertion of CL3 specifically promoted dendritogenesis of cortical neurons by acting upstream of RacGEF STEF and Rac, both present in lipid rafts. Thus, CL3 may represent a key element in the Ca(2+)-dependent and lipid-raft-delineated switch that turns on extrinsic activity-regulated dendrite formation in developing cortical neurons.

Synapsin Phosphorylation by Src Tyrosine Kinase Enhances Src Activity in Synaptic Vesicles

Synapsins are synaptic vesicle-associated phosphoproteins implicated in the regulation of neurotransmitter release. Synapsin I is the major binding protein for the SH3 domain of the kinase c-Src in synaptic vesicles. Its binding leads to stimulation of synaptic vesicle-associated c-Src activity. We investigated the mechanism and role of Src activation by synapsins on synaptic vesicles. We found that synapsin is tyrosine phosphorylated by c-Src in vitro and on intact synaptic vesicles independently of its phosphorylation state on serine. Mass spectrometry revealed a single major phosphorylation site at Tyr(301), which is highly conserved in all synapsin isoforms and orthologues. Synapsin tyrosine phosphorylation triggered its binding to the SH2 domains of Src or Fyn. However, synapsin selectively activated and was phosphorylated by Src, consistent with the specific enrichment of c-Src in synaptic vesicles over Fyn or n-Src. The activity of Src on synaptic vesicles was controlled by the amount of vesicle-associated synapsin, which is in turn dependent on synapsin serine phosphorylation. Synaptic vesicles depleted of synapsin in vitro or derived from synapsin null mice exhibited greatly reduced Src activity and tyrosine phosphorylation of other synaptic vesicle proteins. Disruption of the Src-synapsin interaction by internalization of either the Src SH3 or SH2 domains into synaptosomes decreased synapsin tyrosine phosphorylation and concomitantly increased neurotransmitter release in response to Ca(2+)-ionophores. We conclude that synapsin is an endogenous substrate and activator of synaptic vesicle-associated c-Src and that regulation of Src activity on synaptic vesicles participates in the regulation of neurotransmitter release by synapsin.

Effect of NMDA receptor antagonist on proliferation of neurospheres from embryonic brain

The N-methyl-D-aspartate (NMDA) receptor, a subtype of ionotropic glutamate receptors, plays an important role in the regulation of neuronal development, learning and memory, and neurodegenerative diseases. NMDA receptor blockade enhances neurogenesis in the hippocampal dentate gyrus in vivo. The effect of NMDA receptor antagonist on proliferation of neural progenitor cells, however, remains to be determined. We investigated changes in the diameter and number of neurospheres derived from the embryonic rat brain after NMDA receptor blockade. Cortical progenitor cells were isolated from gestational day 18 fetal rats according to the Percoll density gradient method. Cultured spheres expressed neural progenitor markers, musashi-1 and nestin. Immunohistochemical analysis demonstrated that cells in Dulbecco's modified Eagle medium/F12 containing 1% fetal bovine serum on day 8 differentiated to MAP-2-positive neurons and GFAP-positive astrocytes. The expression of NR1 and NR2B subunits of the NMDA receptor in neurospheres was detected. Neither brief nor sustained exposure to NMDA altered the diameter and number of neurospheres. Brief exposure to 30 μM MK-801, an NMDA receptor antagonist, decreased the diameter of neurospheres. Sustained exposure to 30 μM MK-801 decreased the diameter and number of neurospheres. Our results provide evidence that MK-801 directly decreased proliferation of neural progenitor cells.

Potentially toxic effects of anaesthetics on the developing central nervous system*

A growing body of experimental evidence suggests that anaesthetics, by influencing GABAergic and glutaminergic neural signalling, can have adverse effects on the developing central nervous system. The biological foundation for this is that gamma-aminobutyric acid and glutamate could act non-synaptically, in addition to their role in neurotransmission in the adult brain, in the regulation of neuronal development in the central nervous system. These neurotransmitters and their receptors are expressed from very early stages of central nervous system development and appear to influence neural progenitor proliferation, cell migration and neuronal differentiation. During the synaptogenetic period, pharmacological blockade of N-methyl-d-aspartate (NMDA)-type glutamate receptors as well as stimulation of GABAA receptors has been reported to be associated with increased apoptosis in the developing brain. Importantly, recent data suggest that even low, non-apoptogenic concentrations of anaesthetics can perturb neuronal dendritic development and thus could potentially lead to impairment of developing neuronal networks. The extrapolation of these experimental observations to clinical practice is of course very difficult and requires extreme caution as differences in drug concentrations and exposure times as well as interspecies variations are all important confounding variables. While clinicians should clearly not withhold anaesthesia based on current animal studies, these observations should urge more laboratory and clinical research to further elucidate this issue.

Retinoic acid and nerve growth factor induce differential regulation of nicotinic acetylcholine receptor subunit expression in SN56 cells

Retinoic acid (RA) and nerve growth factor (NGF) have multiple functions in the regulation of neuronal development. In the present study, we characterized the expression of different nicotinic acetylcholine receptor (nAChR) subtypes in the cholinergic SN56 cell line and investigated the roles of RA and NGF in the expression of choline acetyltransferase (ChAT) and different nAChR subtypes. The nAChR agonist [(3)H]epibatidine was bound to two sites, with apparent affinities of 13 and 380 pM. RT-PCR analysis revealed expression of alpha3, alpha4, alpha5, alpha7, beta2, and beta4 nAChR subunits. RA treatment induced morphological changes, and the mRNA level of ChAT was maximally elevated after 4 days of exposure. The density of [(3)H]epibatidine binding sites and the mRNA and protein level of the alpha3 and beta2 nAChR subunits were also increased by RA-induced differentiation. RA down-regulated the mRNA and protein level of the alpha4 nAChR subunit, whereas no significant change was observed in the mRNA and protein level of the alpha7 nAChR subunit. NGF treatment increased the mRNA and protein level of the alpha3 and beta2 nAChR subunits. No morphological effects of NGF were observed, and the mRNA level of ChAT and mRNA and protein level of the alpha4 and alpha7 nAChR subunits were not significantly altered. Validation was performed with real-time RT-PCR. The present results show that RA and NGF have different effects on the expression of ChAT and the morphology and the expression pattern of different nAChR subunits in cholinergic SN56 cells.

N-methyl- d-aspartate receptor expression during adult neurogenesis in the rat dentate gyrus

N-methyl- d-aspartate (NMDA) receptors play a crucial role in the regulation of neuronal development during embryogenesis and they also regulate the rate of neurogenesis and proliferation in the adult dentate gyrus. However, the mechanism by which they influence these processes is not fully understood. NMDA receptors seem to be functional in hippocampal precursor cells and recently generated granule neurons, although there is no anatomical correlate of these physiological observations. We have analyzed the expression of the NMDA receptor subunits NR1 and NR2B in precursor cells and recently generated granule neurons of the adult rat dentate gyrus, using 5′bromodeoxyuridine, green fluorescent protein–retrovirus and immunohistochemistry. Our results indicate that NR1 and NR2B are expressed in some proliferating cells of the adult subgranular zone. These receptors are absent from transiently amplifying progenitors (type 2–3 cells) but they are found in glial fibrillar acidic protein expressing cells in the subgranular zone, suggesting its presence in bipotential (type-1) precursor cells. NR1 and NR2B are rarely found in granule cells younger than 60 h. By contrast, many granule cells generated 14 days before killing express both NMDA receptor subunits. These results demonstrate that adult hippocampal neurogenesis may be regulated by NMDA receptors present in precursor cells and in differentiating granule neurons, although these receptors are probably not located on synapses. However, an indirect effect through NMDA receptors located in other cell types should not be excluded.

Differential Regulation of AMPA Receptor Subunit Trafficking by Palmitoylation of Two Distinct Sites

Modification of AMPA receptor function is a major mechanism for the regulation of synaptic transmission and underlies several forms of synaptic plasticity. Post-translational palmitoylation is a reversible modification that regulates localization of many proteins. Here, we report that palmitoylation of the AMPA receptor regulates receptor trafficking. All AMPA receptor subunits are palmitoylated on two cysteine residues in their transmembrane domain (TMD) 2 and in their C-terminal region. Palmitoylation on TMD 2 is upregulated by the palmitoyl acyl transferase GODZ and leads to an accumulation of the receptor in the Golgi and a reduction of receptor surface expression. C-terminal palmitoylation decreases interaction of the AMPA receptor with the 4.1N protein and regulates AMPA- and NMDA-induced AMPA receptor internalization. Moreover, depalmitoylation of the receptor is regulated by activation of glutamate receptors. These data suggest that regulated palmitoylation of AMPA receptor subunits modulates receptor trafficking and may be important for synaptic plasticity.

Pathogenesis of Lyme neuroborreliosis: Mitogen-activated protein kinases Erk1, Erk2, and p38 in the response of astrocytes to Borrelia burgdorferi lipoproteins

Lyme borreliosis, which is prevalent both in the US and in Europe, is an infectious disease that may cause local inflammation in numerous organs. We have hypothesized that, as with some neurodegenerative diseases, the pathogenesis of the neurocognitive deficiencies associated with Lyme neuroborreliosis of the central nervous system also has an inflammatory component. Dysregulated production of pro-inflammatory cytokines such as IL-6 and TNF-α can lead to neuronal damage. Mitogen-activated protein kinases (MAPK) play a key role in the regulation of neuronal development, growth, and survival, as well as that of pro-inflammatory cytokine production. As a model, we explored the possibility that MAPK-mediated lipoprotein-induced apoptosis and gliosis of rhesus monkey astrocytes stimulated in vitro. Lipoproteins are the key inflammatory molecule type of Borrelia burgdorferi, the spirochete that causes Lyme disease, and we had previously shown that lipoprotein-induced TNF-α production in astrocytes caused astrocyte apoptosis, and IL-6 enhanced proliferation of these cells. Lipoproteins readily activated p38 and Erk1/2 MAPK, thus enlisting these pathways among the kinase pathways that spirochetes may address as they invade the central nervous system. We also investigated whether specific inhibition of p38 and Erk1/2 MAPK would inhibit TNF-α and IL-6 production and thus astrocyte apoptosis, and proliferation, respectively. Lipoprotein-stimulated IL-6 production was unaffected by the MAPK inhibitors. In contrast, inhibition of both p38 and Erk1/2 significantly diminished TNF-α production, and totally abrogated production of this cytokine when both MAPK pathways were inhibited simultaneously. MAPK inhibition thus may be considered as a strategy to control inflammation and apoptosis in Lyme neuroborreliosis.

Protein palmitoylation: a regulator of neuronal development and function.

Palmitoylation — the post-translational modification of proteins with the lipid palmitate — has emerged as an important mechanism for regulating protein trafficking and function. Classic studies showed that palmitoylation targets many signalling enzymes to specialized lipid microdomains on the cytosolic face of the plasma membrane, thereby directing their integration into specific transduction pathways. More recent work shows that palmitate reversibly modifies numerous classes of neuronal proteins, including neurotransmitter receptors, synaptic scaffolding proteins and secreted signalling molecules. This review highlights recent evidence that protein palmitoylation regulates trafficking and signalling pathways that are important for brain development and synaptic transmission.

Expression of neurotrophin messenger RNAs during rat urinary bladder development

The family of neurotrophins, encompassing nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4/5 (NT-4/5), is important in the regulation of neuronal development and function. We examined the expression of neurotrophin messenger RNAs (mRNAs) in the rat urinary bladder during pre- and postnatal development using competitive reverse transcription-polymerase chain reaction. The mRNA levels showed a biphasic pattern of expression; one peak was at prenatal ages (embryonic day (E)15–E18) and the other peak was at postnatal ages (postnatal day (P)14–P28). NT-4/5, BDNF and NGF mRNA levels were greatest at E15, E16 and E18, respectively. In contrast, NT-3 mRNA levels were significantly highest at P14. These data suggest that neurotrophins are involved in the mechanisms of bladder nerve growth for the prenatal period and reorganization of bladder reflex pathways during the second to the fourth postnatal week.

Aminergic receptors during the development of the human brain: the contribution of in vitro imaging techniques

The development of the human brain is a complex process and, in this regard, the maturation of neurotransmitter systems and their receptors is of special interest. The study of these systems requires methodological approaches with powerful anatomical resolution. In this paper we review the application of visualization procedures to the fine localization, pattern of appearance and functional relevance of monoaminergic receptors in postmortem human brain samples corresponding to different stages of development (fetal, neonatal, infant). Data obtained by using mostly in vitro autoradiography but also in situ hybridization and, very recently, second messenger labeling, are discussed, including the methodological limitations inherent in working with inmature human tissue. From these studies, several conclusions were made. (1) It is possible to visualize, in the human brain with high resolution, the presence of neuroreceptors at early prenatal stages. (2) The anatomical distribution of monoaminergic receptors in the developing human brain is, in general terms, comparable to that found in the adult. (3) During the developmental process, some receptors, which are early and sometimes transiently expressed, play important thophic roles in the regulation of neuronal development: this is the case with the serotonin 5-HT 1A receptors, which attain peak levels of hyperexpression over the hippocampus (dentate gyrus, dendritic areas of CA fields) and the raphe nuclei and show a transient expression in the cerebellum, around the 25 week of gestational age. (4) Different patterns of ontogenetic appearance for human receptors have been identified: dopamine D 2-like (caudate, putamen, nigra) and 5-HT 1A receptors are good examples of prenatal development, while 5-HT 1B sites (basal ganglia, neocortex) present a mainly postnatal pattern of appearance. (5) Neurotransmitter receptors at human fetal stages are already functional from the point of view of transducing response.

The role of retinoid metabolism by alcohol and aldehyde dehydrogenases in differentiation of cultured neuronal cells.

Retinoic acid is a morphogen that plays important roles during fetal growth in specification of head and limb bud development (Hofman and Eichele, 1994), and probably also in regulation of neuronal development (Holder and Maden, 1992). Exposure of the developing fetus to ethanol can cause a range of deformities and neurological abnormalities, leading to conditions known in humans as fetal alcohol syndrome (FAS) and alcohol related neurodevelopmental disorder (ARND) (Sampson et al., 1997). Retinoic acid is formed from retinol (vitamin A) via oxidation first to retinal, catalysed by isozymes of alcohol dehydrogenase (ADH) or the short chain dehydrogenase/reductase family. Retinal is then oxidised to retinoic acid by aldehyde dehydrogenase (A1DH), retinal dehydrogenase or microsomal cytochrome P450 1A1 or 1A2 (Duester, 1996). Retinoic acid has its effect on fetal development through binding to nuclear receptors and forming transcription factors which n regulate gene expression (De Luca, 1991; Chambon, 1996). It has been postulated that one of the mechanisms leading to the development of FAS or ARND may be inhibition of retinoic acid production caused by competitive inhibition of alcohol and/or aldehyde dehydrogenases during ethanol metabolism (Duester, 1991, Deltour et al., 1996). e present study was initiated to see whether cultured neuronal cells could be used a model system in which to study the effects of ethanol and acetaldehyde on retinoid- regulated neuronal cell differentiation. A human neuroblastoma cell line, SH-SY5Y, was chosen because it had previously been shown to be responsive to retinoic acid (Pahlman et al., 1984).

Inhibition of dopamine and choline acetyltransferase concentrations in rat CNS neurons by rat alpha 1- and alpha 2-macroglobulins.

Previous studies have implicated human alpha-2-macroglobulin (alpha2M) as a potential regulator of neuronal development and function. Rat alpha-1-macroglobulin (alpha1M) and acute-phase alpha-2-macroglobulin (alpha2M) are murine homologues of human alpha2M. In this report, we tested the effect of intracranially infused serotonin-activated rat alpha1M (5HT-alpha1M) on the concentration of dopamine (DA) in the corpus striatum in vivo and the effect of 5HT-activated rat alpha1M and alpha2M on the choline acetyltransferase (ChAT) activity upon embryonic basal forebrain neurons in culture. The results show that direct infusion of 0.65 nmole rat 5HT-alpha1M into the adult rat corpus striatum produced a consistent attenuation upon striatal DA concentrations. This decrease was particularly prominent at 5-7 days post-infusion. In addition, rat 5HT-alpha1M and rat 5HT-alpha2M, like human 5HT-alpha2M, all significantly inhibited ChAT activity of embryonic rat cerebral cortex neurons. Although normal human alpha2M and rat alpha2M were either marginally or insignificantly inhibitory in this preparation, normal rat alpha1M dose-dependently inhibited ChAT activity. These results demonstrate that monoamine-activated alpha-macroglobulins from rat depress dopaminergic and cholinergic neurotransmitter systems in the CNS, and this suggests a potential regulatory role of these alpha-macroglobulins in neurotransmitter metabolism.