Thyroid hormones (THs) are essential for establishing layered brain structures, a process called corticogenesis, by acting on transcriptional activity of numerous genes. In humans, deficiency of the monocarboxylate transporter 8 (MCT8), involved in cellular uptake of THs before their action, results in severe neurological abnormalities, known as the Allan-Herndon-Dudley syndrome. While the brain lesions predominantly originate prenatally, it remains unclear how and when exactly MCT8 dysfunction affects cellular processes crucial for corticogenesis. We investigated this by inducing in vivo RNAi vector-based knockdown of MCT8 in neural progenitors of the chicken optic tectum, a layered structure that shares many developmental features with the mammalian cerebral cortex. MCT8 knockdown resulted in cellular hypoplasia and a thinner optic tectum. This could be traced back to disrupted cell-cycle kinetics and a premature shift to asymmetric cell divisions impairing progenitor cell pool expansion. Birth-dating experiments confirmed diminished neurogenesis in the MCT8-deficient cell population as well as aberrant migration of both early-born and late-born neuroblasts, which could be linked to reduced reelin signaling and disorganized radial glial cell fibers. Impaired neurogenesis resulted in a reduced number of glutamatergic and GABAergic neurons, but the latter additionally showed decreased differentiation. Moreover, an accompanying reduction in untransfected GABAergic neurons suggests hampered intercellular communication. These results indicate that MCT8-dependent TH uptake in the neural progenitors is essential for early events in corticogenesis, and help to understand the origin of the problems in cortical development and function in Allan-Herndon-Dudley syndrome patients.SIGNIFICANCE STATEMENT Thyroid hormones (THs) are essential to establish the stereotypical layered structure of the human forebrain during embryonic development. Before their action on gene expression, THs require cellular uptake, a process facilitated by the TH transporter monocarboxylate transporter 8 (MCT8). We investigated how and when dysfunctional MCT8 can induce brain lesions associated with the Allan-Herndon-Dudley syndrome, characterized by psychomotor retardation. We used the layered chicken optic tectum to model cortical development, and induced MCT8 deficiency in neural progenitors. Impaired cell proliferation, migration, and differentiation resulted in an underdeveloped optic tectum and a severe reduction in nerve cells. Our data underline the need for MCT8-dependent TH uptake in neural progenitors and stress the importance of local TH action in early development.

Intrabrain transplantation of chromaffin cell aggregates of the Zuckerkandl's organ, an extra-adrenal paraganglion that has never been tested for antiparkinsonian treatment, induced gradual improvement of functional deficits in parkinsonian rats. These beneficial effects were related to long survival of grafted cells, striatal reinnervation, and enhancement of dopamine levels in grafted striatum. Grafted cells were not dopaminergics, but they expressed glial cell line-derived neurotrophic factor (GDNF) and transforming growth factor-beta(1). These factors were detected in the host striatal tissue, indicating that chromaffin cells secreted them after grafting. Because glial cell line-derived neurotrophic factor possesses neurorestorative properties over dopaminergic neurons, and transforming growth factor-beta(1) is a cofactor that potentiates the neurotrophic actions of GDNF, functional regeneration was likely caused by the chronic trophic action of neurotrophic factors delivered by long-surviving grafted cells. This work should stimulate research on the clinical applicability of transplants of the Zuckerkandl's organ in Parkinson's disease.

Adrenergic agents modulate the activity of midbrain ventral tegmental area (VTA) neurons. However, the sources of noradrenergic and adrenergic inputs are not well characterized. Immunostaining for dopamine beta-hydroxylase revealed fibers within dopamine (DA) neuron areas, with the highest density in the retrorubral field (A8 cell group), followed by the VTA (A10 cell group), and very few fibers within substantia nigra compacta. A less dense, but a similar pattern of fibers was also found for the epinephrine marker, phenylethanolamine N-methyl transferase. Injection of the retrograde tracer wheat germ agglutinin-apo (inactivated) horseradish peroxidase conjugated to colloidal gold, or cholera toxin subunit b, revealed that the noradrenergic innervation of the A10 and A8 regions arise primarily from A1, A2, A5, and locus ceruleus neurons. Selective lesions of the ventral noradrenergic bundle confirmed a prominent innervation from A1 and A2 areas. Retrogradely labeled epinephrine neurons were found mainly in the C1 area. The identification of medullary noradrenergic and adrenergic afferents to DA neuron areas indicates new pathways for visceral-related inputs to reward-related areas in the midbrain.

Ovulation disorders are a serious problem for humans and livestock. In female rodents, kisspeptin neurons in the anteroventral periventricular nucleus (AVPV) are responsible for generating a luteinizing hormone (LH) surge and consequent ovulation. Here, we report that adenosine 5-triphosphate (ATP), a purinergic receptor ligand, is a possible neurotransmitter that stimulates AVPV kisspeptin neurons to induce an LH surge and consequent ovulation in rodents. Administration of an ATP receptor antagonist (PPADS) into the AVPV blocked the LH surge in ovariectomized (OVX) rats treated with a proestrous level of estrogen (OVX + high E2) and significantly reduced the ovulation rate in proestrous ovary-intact rats. AVPV ATP administration induced a surge-like LH increase in OVX + high E2 rats in the morning. Importantly, AVPV ATP administration could not induce the LH increase in Kiss1 KO rats. Furthermore, ATP significantly increased intracellular Ca2+ levels in immortalized kisspeptin neuronal cell line, and coadministration of PPADS blocked the ATP-induced Ca2+ increase. Histologic analysis revealed that the proestrous level of estrogen significantly increased the number of P2X2 receptor (an ATP receptor)-immunopositive AVPV kisspeptin neurons visualized by tdTomato in Kiss1-tdTomato rats. The proestrous level of estrogen significantly increased varicosity-like vesicular nucleotide transporter (a purinergic marker)-immunopositive fibers projecting to the vicinity of AVPV kisspeptin neurons. Furthermore, we found that some hindbrain vesicular nucleotide transporter-positive neurons projected to the AVPV and expressed estrogen receptor α, and the neurons were activated by the high E2 treatment. These results suggest that hindbrain ATP-purinergic signaling triggers ovulation via activation of AVPV kisspeptin neurons.SIGNIFICANCE STATEMENT Ovulation disorders, which cause infertility and low pregnancy rates, are a serious problem for humans and livestock. The present study provides evidence that adenosine 5-triphosphate, acting as a neurotransmitter in the brain, stimulates kisspeptin neurons in the anteroventral periventricular nucleus, known as the gonadotropin-releasing hormone surge generator, via purinergic receptors to induce the gonadotropin-releasing hormone/luteinizing hormone surge and ovulation in rats. In addition, histologic analyses indicate that adenosine 5-triphosphate is likely to be originated from the purinergic neurons in the A1 and A2 of the hindbrain. These findings may contribute to new therapeutic controls for hypothalamic ovulation disorders in humans and livestock.

Previous data have strongly implicated hindbrain catecholamine/neuropeptide Y (NPY) coexpressing neurons as key mediators of the glucoprivic feeding response. Catecholamine/NPY cell bodies are concentrated in the A1 and caudal C1 cell cluster (A1/C1) in the ventrolateral medulla, a region highly sensitive to glucoprivic challenge. To further investigate the importance of this catecholamine subpopulation in glucoregulation, we used small interfering RNA (siRNA) technology to produce a targeted gene knockdown of NPY and dopamine-beta-hydroxylase (DBH), a catecholamine biosynthetic enzyme. Unilateral injection of NPY siRNA and DBH siRNA (0.02 nmol each) both significantly inhibited expression of the targeted genes up to 2 d, as revealed by real-time PCR, and reduced protein expression up to 8 d, as revealed by immunohistochemistry, compared with the control nontargeting siRNA (ntRNA) side. Subsequently, targeted siRNA or control ntRNA was injected bilaterally into A1/C1 and responses to 2-deoxy-D-glucose (2DG; 200 mg/kg)-induced glucoprivation were tested 3-7 d later. Silencing of either Npy or Dbh alone did not reduce glucoprivic feeding or hyperglycemic responses, compared with responses of ntRNA-injected controls. In contrast, simultaneous silencing of both Npy and Dbh reduced 2DG-induced feeding by 61%. Neither the hyperglycemic response to 2DG nor feeding elicited by mercaptoacetate (68 mg/kg)-induced blockade of fatty acid oxidation ("lipoprivic feeding") was reduced by simultaneous silencing of these two genes. These results suggest that catecholamines and NPY act conjointly to control glucoprivic feeding and that the crucial NPY/catecholamine coexpressing neurons are concentrated in the A1/C1 cell group.