Comparative analysis of the sea urchin genome has broad implications for the primitive state of deuterostome host defense and the genetic underpinnings of immunity in vertebrates. The sea urchin has an unprecedented complexity of innate immune recognition receptors relative to other animal species yet characterized. These receptor genes include a vast repertoire of 222 Toll-like receptors, a superfamily of more than 200 NACHT domain-leucine-rich repeat proteins (similar to nucleotide-binding and oligomerization domain (NOD) and NALP proteins of vertebrates), and a large family of scavenger receptor cysteine-rich proteins. More typical numbers of genes encode other immune recognition factors. Homologs of important immune and hematopoietic regulators, many of which have previously been identified only from chordates, as well as genes that are critical in adaptive immunity of jawed vertebrates, also are present. The findings serve to underscore the dynamic utilization of receptors and the complexity of immune recognition that may be basal for deuterostomes and predicts features of the ancestral bilaterian form.

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The promoter regions of the rat corticotropin-releasing factor (CRF), oxytocin (OT), and vasopressin (AVP) genes contain sequences similar to the cis-acting response element identified for NGFI-B, an immediate-early gene structurally related to the steroid hormone receptor superfamily. Combined immuno- and hybridization histochemical approaches were used to determine whether challenges that influence the synthesis and secretion of CRF, OT, and/or AVP result in altered expression in neurosecretory neurons of NGFI-B and another immediate-early gene, c-fos, which is widely used as a marker for functionally activated neurons. NGFI-B mRNA was found to be expressed at constitutively high levels in the telencephalon, but not in the endocrine hypothalamus, of unperturbed controls; basal levels of c-fos expression were uniformly low throughout the CNS. NGFI-B and c-fos mRNAs, and Fos protein, were induced with a similar time course and in similar neuroendocrine cell types in response to acute hypotensive hemorrhage (15% reduction in blood volume), intravenous injection of interleukin-1 beta (IL-1 beta; 1.87 micrograms/kg), chronic salt loading (7 d maintenance on 2% saline), and acute bilateral adrenalectomy. c-fos mRNA and Fos protein were readily demonstrable in afferent pathways that have been implicated as mediating the neuroendocrine responses in the three stress paradigms; these include medullary catecholaminergic cell groups in response to IL-1 beta and hemorrhage, and cell groups lining the lamina terminalis in response to salt loading. Challenge-specific induction of NGFI-B expression was detectable in these extrahypothalamic cell groups, though with a lesser sensitivity than that required to reveal NGFI-B induction in the hypothalamus, or c-fos expression in these related afferents. These results establish NGFI-B as a useful adjunct to c-fos, for revealing synaptic and/or transcriptional activation in the magno- and parvocellular neurosecretory systems. Differences in the sensitivity of the two markers in revealing functionally related activation in extrahypothalamic regions speak to general issues concerning the use of immediate-early genes in mapping functional circuitry in the CNS.

Cellular mechanisms underlying learning and memory were investigated in the octopus using a brain slice preparation of the vertical lobe, an area of the octopus brain involved in learning and memory. Field potential recordings revealed long-term potentiation (LTP) of glutamatergic synaptic field potentials similar to that in vertebrates. These findings suggest that convergent evolution has led to the selection of similar activity-dependent synaptic processes that mediate complex forms of learning and memory in vertebrates and invertebrates.

BackgroundRhythmic motor patterns for locomotion in vertebrates are generated in spinal cord neural networks known as spinal Central Pattern Generators (CPGs). A key element in pattern generation is the role of glycinergic synaptic transmission by interneurons that cross the cord midline and inhibit contralaterally-located excitatory neurons. The glycinergic inhibitory drive permits alternating and precisely timed motor output during locomotion such as walking or swimming. To understand better the evolution of this system we examined the physiology of the neural network controlling swimming in an invertebrate chordate relative of vertebrates, the ascidian larva Ciona intestinalis.ResultsA reduced preparation of the larva consisting of nerve cord and motor ganglion generates alternating swimming movements. Pharmacological and genetic manipulation of glycine receptors shows that they are implicated in the control of these locomotory movements. Morphological molecular techniques and heterologous expression experiments revealed that glycine receptors are inhibitory and are present on both motoneurones and locomotory muscle while putative glycinergic interneurons were identified in the nerve cord by labeling with an anti-glycine antibody.ConclusionsIn Ciona intestinalis, glycine receptors, glycinergic transmission and putative glycinergic interneurons, have a key role in coordinating swimming movements through a simple CPG that is present in the motor ganglion and nerve cord. Thus, the strong association between glycine receptors and vertebrate locomotory networks may now be extended to include the phylum chordata. The results suggest that the basic network for 'spinal-like' locomotion is likely to have existed in the common ancestor of extant chordates some 650 M years ago.

The deduced structure of the rat melanin-concentrating hormone (MCH) precursor predicted the existence of at least two peptides that may be processed from it, one similar to teleost MCH and a second novel neuropeptide, NEI. Cellular localization studies confirmed that prepro-MCH (ppMCH) mRNA and the MCH and NEI peptides are expressed predominantly in cells in the zona incerta and caudal lateral hypothalamic area with minor contingents seen in the olfactory tubercle and pons. A moderate MCHand NEl-immunoreactive axonal projection to the median eminence and, particularly, to oxytocin-rich regions of the posterior pituitary suggested some anatomical heterogeneity of ppMCH-expressing neurons in the hypothalamus, and an involvement in neuroendocrine function. In the present study, immunohistochemical and hybridization histochemical methods were used to follow MCH gene and peptide expression as a function of reproductive status in female rats. Nursing dams sacrificed after 8 to 21 days of lactation consistently displayed ppMCH mRNA and MCH and NEI immunoreactivity in a discrete and contiguous band, encompassing the ventral aspect of the medial part of the medial preoptic nucleus, the periventricular preoptic nucleus, and the most rostra1 aspects of the paraventricular nucleus of the hypothalamus (PVH). Combined immunohistochemical (for oxytocin) and hybridization histochemical (for ppMCH mRNA) staining failed to reveal a significant degree of congruence in the two chemicallyspecified cell populations in the PVH of lactating dams. The apparent induction of ppMCH-derived peptides and mRNA in the preoptic area and PVH was not apparent in animals sacrificed 4 to 8 days after weaning, during late pregnancy, or at any point in the estrous cycle. Moreover, no frank alterations in ppMCH mRNA were evident in the principal sites at which ppMCH is expressed at constitutively high levels, i.e. in the lateral hypothalamic area and zona incerta, as a function of reproductive status. The loci and apparent state-dependency of the induction of ppMCH mRNA and peptide expression suggests a role for these gene products in the control of lactation.Melanin-concentrating hormone (MCH) is a cyclic heptadecapeptide first isolatcd from salmon pituitary, which plays a role in effecting adaptive changes in pigmentation in lower vertebrates ( I , 2). Recent cloning of cDNAs encoding the rat hypothalamic MCH precursor predicted the existence of a C-terminal nonadecapcptide (rMCH) which is structurally similar to salmon M C H (sMCH) (3-6). In addition, rat prepro-MCH (ppMCH) appears to encode at least one additional neuropeptide, named, by convention ( 7 ) neuropeptide EI, or NEI (3). Evidence obtained in primary culturcs of dispersed hypothalamic cells indicates that both peptides are indeed processed from ppMCH (8).In the rat, cells expressing ppMCH-derived peptides and mRNA are localized principally in the lateral hypothalamic area and zona incerta, with minor accumulations seen in the olfactory tubercle and pons (9). Fiber systems stained for MC...

To examine the role of the amino acid GABA in the locomotion of basal chordates, we investigated the pharmacology of swimming and the morphology of GABA-immunopositive neurones in tadpole larvae of the ascidians Ciona intestinalis and Ciona savignyi. We verified that electrical recording from the tail reflects alternating muscle activity during swimming by correlating electrical signals with tail beats using high-speed video recording. GABA reversibly reduced swimming periods to single tail twitches, while picrotoxin increased the frequency and duration of electrical activity associated with spontaneous swimming periods. Immunocytochemistry for GABA revealed extensive labelling throughout the larval central nervous system. Two strongly labelled regions on either side of the sensory vesicle were connected by an arc of labelled fibres, from which fibre tracts extended caudally into the visceral ganglion. Fibre tracts extended ventrally from a third, more medial region in the posterior sensory vesicle. Two rows of immunoreactive cell bodies in the visceral ganglion extended neurites into the nerve cord, where varicosities were seen. Thus, presumed GABAergic neurones form a network that could release GABA during swimming that is involved in modulating the time course and frequency of periods of spontaneous swimming. GABAergic and motor neurones in the visceral ganglion could interact at the level of their cell bodies and/or through the presumed GABAergic fibres that enter the nerve cord. The larval swimming network appears to possess some of the properties of spinal networks in vertebrates, while at the same time possibly showing a type of peripheral innervation resembling that in some protostomes.

BackgroundThe retina of craniates/vertebrates has been proposed to derive from a photoreceptor prosencephalic territory in ancestral chordates, but the evolutionary origin of the different cell types making the retina is disputed. Except for photoreceptors, the existence of homologs of retinal cells remains uncertain outside vertebrates.MethodsThe expression of genes expressed in the sensory vesicle of the ascidian Ciona intestinalis including those encoding components of the monoaminergic neurotransmission systems, was analyzed by in situ hybridization or in vivo transfection of the corresponding regulatory elements driving fluorescent reporters. Modulation of photic responses by monoamines was studied by electrophysiology combined with pharmacological treatments.ResultsWe show that many molecular characteristics of dopamine-synthesizing cells located in the vicinity of photoreceptors in the sensory vesicle of the ascidian Ciona intestinalis are similar to those of amacrine dopamine cells of the vertebrate retina. The ascidian dopamine cells share with vertebrate amacrine cells the expression of the key-transcription factor Ptf1a, as well as that of dopamine-synthesizing enzymes. Surprisingly, the ascidian dopamine cells accumulate serotonin via a functional serotonin transporter, as some amacrine cells also do. Moreover, dopamine cells located in the vicinity of the photoreceptors modulate the light-off induced swimming behavior of ascidian larvae by acting on alpha2-like receptors, instead of dopamine receptors, supporting a role in the modulation of the photic response. These cells are located in a territory of the ascidian sensory vesicle expressing genes found both in the retina and the hypothalamus of vertebrates (six3/6, Rx, meis, pax6, visual cycle proteins).ConclusionWe propose that the dopamine cells of the ascidian larva derive from an ancestral multifunctional cell population located in the periventricular, photoreceptive field of the anterior neural tube of chordates, which also gives rise to both anterior hypothalamus and the retina in craniates/vertebrates. It also shows that the existence of multiple cell types associated with photic responses predates the formation of the vertebrate retina.