Transgenerational transmission of aspartame-induced anxiety and changes in glutamate-GABA signaling and gene expression in the amygdala.

We report the effects of aspartame on anxiety-like behavior, neurotransmitter signaling and gene expression in the amygdala, a brain region associated with the regulation of anxiety and fear responses. C57BL/6 mice consumed drinking water containing 0.015% or 0.03% aspartame, a dose equivalent of 8 to 15% of the FDA recommended maximum human daily intake, or plain drinking water. Robust anxiety-like behavior (evaluated using open field test and elevated zero maze) was observed in male and female mice consuming the aspartame-containing water. Diazepam, an allosteric modulator of the GABA-A receptor, alleviated the anxiety-like behavior. RNA sequencing of the amygdala followed by KEGG biological pathway analysis of differentially expressed genes showed glutamatergic and GABAergic synapse pathways as significantly enriched. Quantitative PCR showed upregulation of mRNA for the glutamate NMDA receptor subunit 2D (Grin2d) and metabotropic receptor 4 (Grm4) and downregulation of the GABA-A receptor associated protein (Gabarap) mRNA. Thus, taken together, our diazepam and gene expression data show that aspartame consumption shifted the excitation-inhibition equilibrium in the amygdala toward excitation. Even more strikingly, the anxiety-like behavior, its response to diazepam, and changes in amygdala gene expression were transmitted to male and female offspring in two generations descending from the aspartame-exposed males. Extrapolation of the findings to humans suggests that aspartame consumption at doses below the FDA recommended maximum daily intake may produce neurobehavioral changes in aspartame-consuming individuals and their descendants. Thus, human population at risk of aspartame's potential mental health effects may be larger than current expectations, which only include aspartame-consuming individuals.

Hippocampal neuroinflammation following combined exposure to cyclophosphamide and naproxen in ovariectomized mice.

Aim: Cancer patients undergoing chemotherapy report cognitive changes collectively termed "chemo brain." Neuroinflammation is among the factors believed to contribute to "chemo brain" suggesting a potential beneficial role for anti-inflammatory drugs in cancer patients undergoing chemotherapy. We investigated whether the non-steroidal anti-inflammatory drug naproxen influenced hippocampal inflammation in non-tumor bearing female mice receiving the chemotherapy drug cyclophosphamide (CP).Materials and methods: Intact and ovariectomized C57BL/6 mice were used to examine potential role of ovarian hormones on neuroinflammation. The mice were placed on naproxen (375 ppm) or control diet, and a week later CP (100 mg/kg; i.p.) was administered every 3 days for 2 weeks. We analyzed hippocampal inflammatory biomarkers, anxiety-like behavior, spatial working memory, exploratory behavior, spontaneous locomotor activity and depression-like behavior.Results: CP produced significant effects on anti-inflammatory but not pro-inflammatory biomarkers. However, CP and naproxen in combination produced significant effects on both pro- and anti- inflammatory biomarkers. Naproxen and ovariectomy individually produced significant effects on pro- and anti-inflammatory biomarkers as well. Working memory and depression-like behavior were not significantly influenced by CP, naproxen or ovariectomy individually although CP and ovariectomy produced significant interaction effects on depression-like behavior. Exploratory behavior and locomotor activity showed significant effects of CP, and interaction between CP and naproxen was significant for locomotor activity.Conclusions: Ovariectomy, naproxen and a combination of CP and naproxen upregulate hippocampal pro- and anti- inflammatory biomarkers. None of the factors individually produce significant behavioral changes that could be consistent with chemo brain, although CP and ovariectomy in combination produced significant effects on depression-like behavior, a co-morbidity of chemo brain.

Frontal Cortical Monoamine Release, Attention, and Working Memory in a Perinatal Nicotine Exposure Mouse Model Following Kappa Opioid Receptor Antagonism.

Perinatal nicotine exposure (PNE) produces frontal cortical hypo-dopaminergic state and attention and working memory deficits consistent with neurodevelopmental disorders such as attention deficit hyperactivity disorder (ADHD). Methylphenidate alleviates ADHD symptoms by increasing extracellular dopamine and noradrenaline. Kappa opioid receptor (KOR) antagonism may be another mechanism to achieve the same results because KOR activation inhibits frontal cortical dopamine release. We administered the selective KOR antagonist norbinaltorphimine (norBNI) (20 mg/kg; intraperitoneal) or methylphenidate (0.75 mg/kg; intraperitoneal) to PNE mouse model and examined frontal cortical monoamine release, attention, and working memory. Both compounds increased dopamine and noradrenaline release but neither influenced serotonin release. Both compounds improved object-based attention and working memory in the PNE group, with norBNI's effects evident at 2.5 h and 5.5 h but absent at 24 h. Methylphenidate's effects were evident at 0.5 h but not at 2.5 h. norBNI's effects temporally coincided with frontal cortical c-Jun N-terminal kinase phosphorylation. norBNI did not alter tissue dopamine content in the nucleus accumbens, offering preliminary support for lack of reinforcement.

Repetitive Mild Traumatic Brain Injury in a Perinatal Nicotine Exposure Mouse Model of Attention Deficit Hyperactivity Disorder.

Attention deficit hyperactivity disorder (ADHD) increases the risk for concussion or mild traumatic brain injury (mTBI). At the same time, recommendations for the management of ADHD include participation in sports and other organized physical activities, including those that carry an increased risk of mTBI. Very little work has been done to determine the extent to which untreated ADHD adversely impacts behavioral outcomes of repeated mild concussions. Here, we used a perinatal nicotine exposure (PNE) mouse model of ADHD combined with a closed-head, repetitive mTBI model. The PNE mouse model carries significant construct, face, and predictive validity as a preclinical model of ADHD. Two-month-old PNE and control mice were subjected to closed-head repetitive mTBI or sham procedure once daily for 5 days. Object-based attention, novel object recognition memory, spatial working memory, and depression-like behavior were analyzed 1 day and 2 weeks following repeated mTBI. Consistent with our previous reports, mice in the PNE group showed significant deficits in object-based attention and working memory prior to mTBI. These deficits persisted following the repeated mTBI. Repeated mTBI produced a transient attention deficit in the control group but did not exacerbate the attention deficit that is characteristic of the PNE group. Although neither PNE nor repetitive mTBI alone influenced immobility in the tail suspension test, when PNE mice were subjected to mTBI, there was a transient increase in this measurement suggesting a synergistic effect of ADHD and mTBI on depression-like behavior. Thus, our data using the PNE mouse model suggest that ADHD may be a risk factor for transient depression following repeated mTBI and that repeated mTBI may be a risk factor for transient attention deficit.

Heritable consequences of paternal nicotine exposure: from phenomena to mechanisms†.

Our understanding of the interactions between genetic and environmental factors in shaping behavioral phenotypes has expanded to include environment-induced epigenetic modifications and the intriguing possibility of their association with heritable behavioral phenotypes. The molecular basis of heritability of phenotypes arising from environment-induced epigenetic modifications is not well defined yet. However, phenomenological evidence in favor of it is accumulating rapidly. The resurgence of interest has led to focus on epigenetic modification of germ cells as a plausible mechanism of heritability. Perhaps partly because of practical reasons such as ease of access to male germ cells compared to female germ cells, attention has turned toward heritable effects of environmental influences on male founders. Public health implications of heritable effects of paternal exposures to addictive substances or to psycho-social factors may be enormous. Considering nicotine alone, over a billion people worldwide use nicotine-containing products, and the majority are men. Historically, the adverse effects of nicotine use by pregnant women received much attention by scientists and public policy experts alike. The implications of nicotine use by men for the physical and mental well-being of their children were not at the forefront of research until recently. Here, we review progress in the emerging field of heritable effects of paternal nicotine exposure and its implications for behavioral health of individuals in multiple generations.

Nicotine exposure of male mice produces behavioral impairment in multiple generations of descendants.

Use of tobacco products is injurious to health in men and women. However, tobacco use by pregnant women receives greater scrutiny because it can also compromise the health of future generations. More men smoke cigarettes than women. Yet the impact of nicotine use by men upon their descendants has not been as widely scrutinized. We exposed male C57BL/6 mice to nicotine (200 µg/mL in drinking water) for 12 wk and bred the mice with drug-naïve females to produce the F1 generation. Male and female F1 mice were bred with drug-naïve partners to produce the F2 generation. We analyzed spontaneous locomotor activity, working memory, attention, and reversal learning in male and female F1 and F2 mice. Both male and female F1 mice derived from the nicotine-exposed males showed significant increases in spontaneous locomotor activity and significant deficits in reversal learning. The male F1 mice also showed significant deficits in attention, brain monoamine content, and dopamine receptor mRNA expression. Examination of the F2 generation showed that male F2 mice derived from paternally nicotine-exposed female F1 mice had significant deficits in reversal learning. Analysis of epigenetic changes in the spermatozoa of the nicotine-exposed male founders (F0) showed significant changes in global DNA methylation and DNA methylation at promoter regions of the dopamine D2 receptor gene. Our findings show that nicotine exposure of male mice produces behavioral changes in multiple generations of descendants. Nicotine-induced changes in spermatozoal DNA methylation are a plausible mechanism for the transgenerational transmission of the phenotypes. These findings underscore the need to enlarge the current focus of research and public policy targeting nicotine exposure of pregnant mothers by a more equitable focus on nicotine exposure of the mother and the father.

Effects of Developmental Nicotine Exposure on Frontal Cortical GABA-to-Non-GABA Neuron Ratio and Novelty-Seeking Behavior.

Cigarette smoking during pregnancy is a major public health concern, resulting in detrimental health effects in the mother and her offspring. The adverse behavioral consequences for children include increased risk for attention deficit hyperactivity disorder, working memory deficits, epilepsy, novelty-seeking, and risk-taking behaviors. Some of these behavioral conditions are consistent with an imbalance in frontal cortical excitatory (glutamate) and inhibitory (GABA) neurotransmitter signaling. We used a GAD67-GFP knock-in mouse model to examine if developmental nicotine exposure alters frontal cortical GABA neuron numbers, GABA-to-non-GABA neuron ratio and behavioral phenotypes. Female mice were exposed to nicotine (100 or 200 µg/mL) in drinking water beginning 3 weeks prior to breeding and until 3 weeks postpartum. Male and female offspring were examined beginning at 60 days of age. The nicotine exposure produced dose-dependent decreases in GABA-to-non-GABA neuron ratios in the prefrontal and medial prefrontal cortices without perturbing the intrinsic differences in cortical thickness and laminar distribution of GABA or non-GABA neurons between these regions. A significant increase in exploratory behavior and a shift toward "approach" in the approach-avoidance paradigm were also observed. Thus, developmental nicotine exposure shifts the cortical excitation-inhibition balance toward excitation and produces behavioral changes consistent with novelty-seeking behavior.

Translational effects and coding potential of an upstream open reading frame associated with DOPA Responsive Dystonia.

Upstream open reading frames (uORFs) have emerged as major post-transcriptional regulatory elements in eukaryotic species. In general, uORFs are initiated by a translation start codon within the 5' untranslated region of a gene (upstream ATG; uATG), and they are negatively correlated with translational efficiency. In addition to their translational regulatory role, some uORFs can code for biologically active short peptides. The importance of uATGs/uORFs is further underscored by human diseases associated with single nucleotide polymorphisms (SNPs), which disrupt existing uORFs or introduce novel uORFs. Although several functional proteins translated from naturally occurring uORFs have been described, the coding potential of uORFs created by SNPs has been ignored because of the a priori assumption that these proteins are short-lived with no likely impact on protein homeostasis. Thus, studies on SNP-created uORFs are limited to their translational effects, leaving unexplored the potential cellular consequences of a SNP/uORF-encoded protein. Here, we investigate functionality of a uATG/uORF introduced by a +142C>T SNP within the GCH1 gene and associated with a familial form of DOPA Responsive Dystonia. We report that the +142C>T SNP represses GCH1 translation, and introduces a short, frame shifted uORF that encodes a 73-amino acid peptide. This peptide is localized within the nucleus and compromises cell viability upon proteasome inhibition. Our work extends the list of uATG/uORF associated diseases and advances research on peptides translated from SNP-introduced uORFs, a neglected component of the proteome.

Cocaine-induced neurodevelopmental deficits and underlying mechanisms.

Exposure to drugs early in life has complex and long-lasting implications for brain structure and function. This review summarizes work to date on the immediate and long-term effects of prenatal exposure to cocaine. In utero cocaine exposure produces disruptions in brain monoamines, particularly dopamine, during sensitive periods of brain development, and leads to permanent changes in specific brain circuits, molecules, and behavior. Here, we integrate clinical studies and significance with mechanistic preclinical studies, to define our current knowledge base and identify gaps for future investigation. Birth Defects Research (Part C) 108:147-173, 2016. © 2016 Wiley Periodicals, Inc.

Prenatal Cocaine Exposure Alters BDNF-TrkB Signaling in the Embryonic and Adult Brain.

Prenatal cocaine exposure remains a major public health concern because of its adverse effects on cognitive function. Although the molecular mechanisms underlying the cognitive impairment are not fully understood, brain-derived neurotrophic factor (BDNF) signaling via its receptor tyrosine kinase B (TrkB) is emerging as a potential candidate. We used a mouse model to examine the impact of ongoing cocaine exposure on BDNF expression in the dorsal forebrain on embryonic day 15 (E15) as well as the long-term effects of prenatal cocaine exposure on BDNF-TrkB signaling in the frontal cortex in early postnatal (postnatal day 16; P16) and adult (P60) male and female mice. We found that ongoing cocaine exposure decreased BDNF expression in the E15 dorsal forebrain, prenatal cocaine exposure did not alter BDNF or TrkB (total or phosphorylated) expression in the frontal cortex at P16, and that the prenatal cocaine exposure produced significant increases in BDNF, the activated (phosphorylated) form of TrkB, as well as Bdnf mRNA in the frontal cortex at P60. The increase in BDNF protein and mRNA expression at P60 was concurrent with hyperacetylation of histone H3 at the Bdnf promoter in the frontal cortex. The increase in frontal cortical BDNF and activated TrkB at P60 occurred in male but not female mice. Thus, our data demonstrate that ongoing cocaine exposure produces a decrease in BDNF expression in the embryonic brain, and that prenatal cocaine exposure produces a sex-specific increase in frontal cortical BDNF-TrkB signaling at P60 only in male mice. Lastly, hyperacetylation of histone H3 at the Bdnf promoter is one epigenetic mechanism mediating the effects of prenatal cocaine exposure on Bdnf expression at P60 in male mice.

Reversal Learning Deficits Associated with Increased Frontal Cortical Brain-Derived Neurotrophic Factor Tyrosine Kinase B Signaling in a Prenatal Cocaine Exposure Mouse Model.

Prenatal cocaine exposure remains a major public health concern because of its adverse impact on cognitive function in children and adults. We report that prenatal cocaine exposure produces significant deficits in reversal learning, a key component of cognitive flexibility, in a mouse model. We used an olfactory reversal learning paradigm and found that the prenatally cocaine-exposed mice showed a marked failure to learn the reversed paradigm. Because brain-derived neurotrophic factor (BDNF) is a key regulator of cognitive functions, and because prenatal cocaine exposure increases the expression of BDNF and the phosphorylated form of its receptor, tyrosine kinase B (TrkB), we examined whether BDNF-TrkB signaling is involved in mediating the reversal learning deficit in prenatally cocaine-exposed mice. Systemic administration of a selective TrkB receptor antagonist restored normal reversal learning in prenatally cocaine-exposed mice, suggesting that increased BDNF-TrkB signaling may be an underlying mechanism of reversal learning deficits. Our findings provide novel mechanistic insights into the reversal learning phenomenon and may have significant translational implications because impaired cognitive flexibility is a key symptom in psychiatric conditions of developmental onset.

Transgenerational transmission of hyperactivity in a mouse model of ADHD.

Attention deficit hyperactivity disorder (ADHD) is a neurobehavioral disorder affecting children and adults. Genetic and environmental factors are associated with the etiology of ADHD. Among the environmental factors, exposure of the developing brain to nicotine is considered a major risk factor. Recent evidence suggests that environmental influences on the brain and behavior may be transmitted from one generation to the next. We used a prenatal nicotine exposure (PNE) mouse model of ADHD to test the hypothesis that PNE-induced hyperactivity, a proxy for human ADHD phenotype, is transmitted from one generation to the next. Our data reveal transgenerational transmission of PNE-induced hyperactivity in mice via the maternal but not the paternal line of descent. We suggest that transgenerational transmission is a plausible mechanism for propagation of environmentally induced ADHD phenotypes in the population.

Preclinical Models of Attention Deficit Hyperactivity Disorder: Neurobiology, Drug Discovery, and Beyond.

OBJECTIVE: We offer an overview of ADHD research using mouse models of nicotine exposure. METHOD: Nicotine exposure of C57BL/6 or Swiss Webster mice occurred during prenatal period only or during the prenatal and the pre-weaning periods. Behavioral, neuroanatomical and neurotransmitter assays were used to investigate neurobiological mechanisms of ADHD and discover candidate ADHD medications. RESULTS: Our studies show that norbinaltorphimine, a selective kappa opioid receptor antagonist is a candidate novel non-stimulant ADHD treatment and that a combination of methylphenidate and naltrexone has abuse deterrent potential with therapeutic benefits for ADHD. Other studies showed transgenerational transmission of ADHD-associated behavioral traits and demonstrated that interactions between untreated ADHD and repeated mild traumatic brain injury produced behavioral traits not associated with either condition alone. CONCLUSION: Preclinical models contribute to novel insights into ADHD neurobiology and are valuable tools for drug discovery and translation to benefit humans with ADHD.

Prenatal nicotine exposure mouse model showing hyperactivity, reduced cingulate cortex volume, reduced dopamine turnover, and responsiveness to oral methylphenidate treatment.

Cigarette smoking, nicotine replacement therapy, and smokeless tobacco use during pregnancy are associated with cognitive disabilities later in life in children exposed prenatally to nicotine. The disabilities include attention deficit hyperactivity disorder (ADHD) and conduct disorder. However, the structural and neurochemical bases of these cognitive deficits remain unclear. Using a mouse model we show that prenatal nicotine exposure produces hyperactivity, selective decreases in cingulate cortical volume, and radial thickness, as well as decreased dopamine turnover in the frontal cortex. The hyperactivity occurs in both male and female offspring and peaks during the "active" or dark phase of the light/dark cycle. These features of the mouse model closely parallel the human ADHD phenotype, whether or not the ADHD is associated with prenatal nicotine exposure. A single oral, but not intraperitoneal, administration of a therapeutic equivalent dose (0.75 mg/kg) of methylphenidate decreases the hyperactivity and increases the dopamine turnover in the frontal cortex of the prenatally nicotine exposed mice, once again paralleling the therapeutic effects of this compound in ADHD subjects. Collectively, our data suggest that the prenatal nicotine exposure mouse model has striking parallels to the ADHD phenotype not only in behavioral, neuroanatomical, and neurochemical features, but also with respect to responsiveness of the behavioral phenotype to methylphenidate treatment. The behavioral, neurochemical, and anatomical biomarkers in the mouse model could be valuable for evaluating new therapies for ADHD and mechanistic investigations into its etiology.

Learning and memory deficits produced by aspartame are heritable via the paternal lineage.

Environmental exposures produce heritable traits that can linger in the population for one or two generations. Millions of individuals consume substances such as artificial sweeteners daily that are declared safe by regulatory agencies without evaluation of their potential heritable effects. We show that consumption of aspartame, an FDA-approved artificial sweetener, daily for up to 16-weeks at doses equivalent to only 7-15% of the FDA recommended maximum daily intake value (equivalent to 2-4 small, 8 oz diet soda drinks per day) produces significant spatial learning and memory deficits in mice. Moreover, the cognitive deficits are transmitted to male and female descendants along the paternal lineage suggesting that aspartame's adverse cognitive effects are heritable, and that they are more pervasive than current estimates, which consider effects in the directly exposed individuals only. Traditionally, deleterious environmental exposures of pregnant and nursing women are viewed as risk factors for the health of future generations. Environmental exposures of men are not considered to pose similar risks. Our findings suggest that environmental exposures of men can produce adverse impact on cognitive function in future generations and demonstrate the need for considering heritable effects via the paternal lineage as part of the regulatory evaluations of artificial sweeteners.

Cocaine alters BDNF expression and neuronal migration in the embryonic mouse forebrain.

Prenatal cocaine exposure impairs brain development and produces lasting alterations in cognitive function. In a prenatal cocaine exposure mouse model, we found that tangential migration of GABA neurons from the basal to the dorsal forebrain and radial neuron migration within the dorsal forebrain were significantly decreased during the embryonic period. The decrease in the tangential migration occurred early in gestation and normalized by late gestation, despite ongoing cocaine exposure. The decrease in radial migration was associated with altered laminar positioning of neurons in the medial prefrontal cortex. The cocaine exposure led to transient decreases in the expression of Tbr2 and Tbr1, transcription factors associated with intermediate progenitor cells and newborn neurons of the dorsal forebrain, respectively, although neurogenesis was not significantly altered. Since cocaine can modulate brain derived neurotrophic factor (BDNF) expression in the mature brain, we examined whether cocaine can alter BDNF expression in the embryonic brain. We found a transient decrease in BDNF protein expression in the cocaine-exposed embryonic forebrain early in gestation. By late gestation, the BDNF expression recovered to control levels, despite ongoing cocaine exposure. In basal forebrain explants from cocaine-exposed embryos, cell migration was significantly decreased, corroborating the in vivo data on tangential GABA neuron migration. Since BDNF can influence tangential neuronal migration, we added BDNF to the culture medium and observed increased cell migration. Our data suggest that cocaine can alter tangential and radial neuronal migration as well as BDNF expression in the embryonic brain and that decreased BDNF may mediate cocaine's effects on neuronal migration.

Prenatal cocaine exposure decreases parvalbumin-immunoreactive neurons and GABA-to-projection neuron ratio in the medial prefrontal cortex.

Cocaine abuse during pregnancy produces harmful effects not only on the mother but also on the unborn child. The neurotransmitters dopamine and serotonin are known as the principal targets of the action of cocaine in the fetal and postnatal brain. However, recent evidence suggests that cocaine can impair cerebral cortical GABA neuron development and function. We sought to analyze the effects of prenatal cocaine exposure on the number and distribution of GABA and projection neurons (inhibitory interneurons and excitatory output neurons, respectively) in the mouse cerebral cortex. We found that the prenatal cocaine exposure decreased GABA neuron numbers and GABA-to-projection neuron ratio in the medial prefrontal cortex of 60-day-old mice. The neighboring prefrontal cortex did not show significant changes in either of these measures. However, there was a significant increase in projection neuron numbers in the prefrontal cortex but not in the medial prefrontal cortex. Thus, the effects of cocaine on GABA and projection neurons appear to be cortical region specific. The population of parvalbumin-immunoreactive GABA neurons was decreased in the medial prefrontal cortex following the prenatal cocaine exposure. The cocaine exposure also delayed the developmental decline in the volume of the medial prefrontal cortex. Thus, prenatal cocaine exposure produced persisting and region-specific effects on cortical cytoarchitecture and impaired the physiological balance between excitatory and inhibitory neurotransmission. These structural changes may underlie the electrophysiological and behavioral effects of prenatal cocaine exposure observed in animal models and human subjects.

Neurogenesis and neuronal migration in the forebrain of the TorsinA knockout mouse embryo.

Early-onset generalized torsion dystonia, also known as DYT1 dystonia, is a childhood onset heritable neurological movement disorder involving painful, involuntary muscle contractions, sustained abnormal postures, and repetitive movements. It is caused by a GAG deletion in the Tor1A gene located on chromosome 9. TorsinA, the product of the Tor1A gene, is expressed throughout the brain beginning early in embryonic development. It plays a role in the regulation of nuclear envelope-cytoskeletal interactions, and presumably nuclear translocation. Since nuclear translocation, powered by cytoskeletal traction, is critical for cell proliferation and migration, we examined whether neurogenesis and neuronal migration are affected in Tor1A-/- mouse brain. Our data show that interkinetic nuclear migration and the pattern of migration of newly generated neurons are impaired in the dorsal forebrain of the Tor1A-/- embryo. However, neurogenesis is not altered significantly. The rate of migration of cells from explants of the medial ganglionic eminence is also impaired in the Tor1A-/- embryo. Thus, loss of torsinA results in subtle but significant alterations in cell proliferation and migration in the embryonic forebrain. These subtle developmental changes are consistent with a lack of significant changes in neuronal numbers, neuronal positioning or size of brain regions in DYT1 dystonia patients.

Compartment-specific transcription factors orchestrate angiogenesis gradients in the embryonic brain.

Prevailing notions of cerebral vascularization imply that blood vessels sprout passively into the brain parenchyma from pial vascular plexuses to meet metabolic needs of growing neuronal populations. Endothelial cells, building blocks of blood vessels, are thought to be homogeneous in the brain with respect to their origins, gene expression patterns and developmental mechanisms. These current notions that cerebral angiogenesis is regulated by local environmental signals contrast with current models of cell-autonomous regulation of neuronal development. Here we demonstrate that telencephalic angiogenesis in mice progresses in an orderly, ventral-to-dorsal gradient regulated by compartment-specific homeobox transcription factors. Our data offer new perspectives on intrinsic regulation of angiogenesis in the embryonic telencephalon, call for a revision of the current models of telencephalic angiogenesis and support novel roles for endothelial cells in brain development.