Amphioxus neuroglia: Molecular characterization and evidence for early compartmentalization of the developing nerve cord.

Glial cells play important roles in the development and homeostasis of metazoan nervous systems. However, while their involvement in the development and function in the central nervous system (CNS) of vertebrates is increasingly well understood, much less is known about invertebrate glia and the evolutionary history of glial cells more generally. An investigation into amphioxus glia is therefore timely, as this organism is the best living proxy for the last common ancestor of all chordates, and hence provides a window into the role of glial cell development and function at the transition of invertebrates and vertebrates. We report here our findings on amphioxus glia as characterized by molecular probes correlated with anatomical data at the transmission electron microscopy (TEM) level. The results show that amphioxus glial lineages express genes typical of vertebrate astroglia and radial glia, and that they segregate early in development, forming what appears to be a spatially separate cell proliferation zone positioned laterally, between the dorsal and ventral zones of neural cell proliferation. Our study provides strong evidence for the presence of vertebrate-type glial cells in amphioxus, while highlighting the role played by segregated progenitor cell pools in CNS development. There are implications also for our understanding of glial cells in a broader evolutionary context, and insights into patterns of precursor cell deployment in the chordate nerve cord.

Amphioxus neurocircuits, enhanced arousal, and the origin of vertebrate consciousness.

Gene expression studies have recently identified the amphioxus homolog of a domain comprising the combined caudal diencephalon plus midbrain, regions implicated in locomotory control and some forms of primary consciousness in vertebrates. The results of EM-level reconstructions of the larval brain of amphioxus, reviewed here, highlight the importance of inputs to this region for light and physical contact, both of which impinge on the same synaptic zone. The neural circuitry provides a starting point for understanding the organization and evolution of locomotory control and arousal in vertebrates, and implies that one of the tasks of midbrain-based consciousness, as it first emerged in vertebrates, would have been to distinguish between light and physical contact, probably sharp pain in the latter case, by assigning different qualia to each. If so, investigating midbrain circuitry more fully could lead to a better understanding of the neural basis of some forms of sensory experience.

The HMGA gene family in chordates: evolutionary perspectives from amphioxus.

High mobility group A proteins of vertebrates, HMGA1 and 2, are chromatin architectural factors involved in development, cell differentiation, and neoplastic transformation. Here, we characterize an amphioxus HMGA gene ortholog and analyze its expression. As a basal chordate, amphioxus is well placed to provide insights into the evolution of the HMGA gene family, particularly in the transition from invertebrates to vertebrates. Our phylogenetic analysis supports the basal position of amphioxus, echinoderm, and hemichordate HMGA sequences to those of vertebrate HMGA1 and HMGA2. Consistent with this, the genomic landscape around amphioxus HMGA shares features with both. Whole mount in situ hybridization shows that amphioxus HMGA mRNA is detectable from neurula stage onwards in both nervous and non-nervous tissues. This correlates with protein expression monitored immunocytochemically using antibodies against human HMGA2 protein, revealing especially high levels of expression in cells of the lamellar body, the amphioxus homolog of the pineal, suggesting that the gene may have, among its many functions, an evolutionarily conserved role in photoreceptor differentiation.

The Cambrian fossil Pikaia, and the origin of chordate somites.

The Middle Cambrian fossil Pikaia has a regular series of vertical bands that, assuming chordate affinities, can be interpreted as septa positioned between serial myotomes. Whether Pikaia has a notochord and nerve cord is less certain, as the dorsal organ, which has no obvious counterpart in living chordates, is the only clearly defined axial structure extending the length of the body. Without a notochord to serve as a reference point, the location of the nerve cord is then conjectural, which begs the question of how a dorsal neural center devoted to somite innervation would first have arisen from a more diffuse ancestral plexus of intraepithelial nerves. This question is examined using hemichordates as a reference point, first for the information they provide on the organization of the ancestral deuterostome nervous system, and second, extending the analysis of E. E. Ruppert, to explain why neural infoldings like the enteropneust collar cord would first have evolved. Both implicate the medial surface of the anterior-most part of the metacoel as the likely site for the evolution of the first somites. The analysis highlights the importance of the somatobranchial condition in chordates, meaning the linkage between the anterior trunk, hox1 expression, and the beginning of the gill series and somites. This feature is arguably a valid criterion by which to assess extinct taxa from the Cambrian that resemble chordates (e.g., vetulicolians and yunnanozoans), but may be unrelated to them. In a more speculative vein, the nature of the dorsal organ is discussed, including the possibility that it is an expanded neural tube combining neural and support functions in one structure.

Mental causation: an evolutionary perspective.

The relationship between consciousness and individual agency is examined from a bottom-up evolutionary perspective, an approach somewhat different from other ways of dealing with the issue, but one relevant to the question of animal consciousness. Two ways are identified that would decouple the two, allowing consciousness of a limited kind to exist without agency: (1) reflex pathways that incorporate conscious sensations as an intrinsic component (InCs), and (2) reflexes that are consciously conditioned and dependent on synaptic plasticity but not memory (CCRs). Whether InCs and CCRs exist as more than hypothetical constructs is not clear, and InCs are in any case limited to theories where consciousness depends directly on EM field-based effects. Consciousness with agency, as we experience it, then belongs in a third category that allows for deliberate choice of alternative actions (DCs), where the key difference between this and CCR-level pathways is that DCs require access to explicit memory systems whereas CCRs do not. CCRs are nevertheless useful from a heuristic standpoint as a conceptual model for how conscious inputs could act to refine routine behaviors while allowing evolution to optimize phenomenal experience (i.e., qualia) in the absence of individual agency, a somewhat counterintuitive result. However, so long as CCRs are not a required precondition for the evolution of memory-dependent DC-level processes, the later could have evolved first. If so, the adaptive benefit of consciousness when it first evolved may be linked as much to the role it plays in encoding memories as to any other function. The possibility that CCRs are more than a theoretical construct, and have played a role in the evolution of consciousness, argues against theories of consciousness focussed exclusively on higher-order functions as the appropriate way to deal with consciousness as it first evolved, as it develops in the early postnatal period of life, or with the conscious experiences of animals other than ourselves. An evolutionary perspective also resolves the problem of free will, that it is best treated as a property of a species rather than the individuals belonging to that species whereas, in contrast, agency is an attribute of individuals.

Consciousness and its hard problems: separating the ontological from the evolutionary.

Few of the many theories devised to account for consciousness are explicit about the role they ascribe to evolution, and a significant fraction, by their silence on the subject, treat evolutionary processes as being, in effect, irrelevant. This is a problem for biological realists trying to assess the applicability of competing theories of consciousness to taxa other than our own, and across evolutionary time. Here, as an aid to investigating such questions, a consciousness "machine" is employed as conceptual device for thinking about the different ways ontology and evolution contribute to the emergence of a consciousness composed of distinguishable contents. A key issue is the nature of the evolutionary innovations required for any kind of consciousness to exist, specifically whether this is due to the underappreciated properties of electromagnetic (EM) field effects, as in neurophysical theories, or, for theories where there is no such requirement, including computational and some higher-order theories (here, as a class, algorithmic theories), neural connectivity and the pattern of information flow that connectivity encodes are considered a sufficient explanation for consciousness. In addition, for consciousness to evolve in a non-random way, there must be a link between emerging consciousness and behavior. For the neurophysical case, an EM field-based scenario shows that distinct contents can be produced in the absence of an ability to consciously control action, i.e., without agency. This begs the question of how agency is acquired, which from this analysis would appear to be less of an evolutionary question than a developmental one. Recasting the problem in developmental terms highlights the importance of real-time feedback mechanisms for transferring agency from evolution to the individual, the implication being, for a significant subset of theories, that agency requires a learning process repeated once in each generation. For that subset of theories the question of how an evolved consciousness can exist will then have two components, of accounting for conscious experience as a phenomenon on the one hand, and agency on the other. This reduces one large problem to two, simplifying the task of investigation and providing what may prove an easier route toward their solution.

An evolutionary perspective on chordate brain organization and function: insights from amphioxus, and the problem of sentience.

The similarities between amphioxus and vertebrate brains, in their regional subdivision, cell types and circuitry, make the former a useful benchmark for understanding the evolutionary innovations that shaped the latter. Locomotory control systems were already well developed in basal chordates, with the ventral neuropile of the dien-mesencephalon serving to set levels of activity and initiate locomotory actions. A chief deficit in amphioxus is the absence of complex vertebrate-type sense organs. Hence, much of vertebrate story is one of progressive improvement both to these and to sensory experience more broadly. This has two aspects: (i) anatomical and neurocircuitry innovations in the organs of special sense and the brain centres that process and store their output, and (ii) the emergence of primary consciousness, i.e. sentience. With respect to the latter, a bottom up, evolutionary perspective has a different focus from a top down human-centric one. At issue: the obstacles to the emergence of sentience in the first instance, the sequence of addition of new contents to evolving consciousness, and the homology relationship between them. A further question, and a subject for future investigation, is how subjective experience is optimized for each sensory modality. This article is part of the theme issue 'Systems neuroscience through the lens of evolutionary theory'.

On the origins and evolution of qualia: An experience-space perspective.

This paper elaborates on a proposal for mapping a configuration space for selector circuits (SCs), defined as the subset of neural correlates of consciousness (NCCs) responsible for evoking particular qualia, to its experiential counterpart, experience-space (E-space), as part of an investigation into the nature of conscious experience as it first emerged in evolution. The dimensionality of E-space, meaning the degrees of freedom required to specify the properties of related sets of qualia, is at least two, but the utility of E-space as a hypothetical construct is much enhanced by assuming it is a large dimensional space, with at least several times as many dimensions as there are categories of qualia to occupy them. Phenomenal consciousness can then be represented as having originated as one or more multidimensional ur-experiences that combined multiple forms of experience together. Taking this as a starting point, questions concerning evolutionary sequence can be addressed, including how the quale best suited to a given sensory modality would have been extracted by evolution from a larger set of possibilities, a process referred to here as dimensional sorting, and how phenomenal consciousness would have been experienced in its earliest manifestations. There is a further question as to whether the E-space formulation is meaningful in analytical terms or simply a descriptive device in graphical form, but in either case it provides a more systematic way of thinking about early stages in the evolution of consciousness than relying on narrative and conjecture alone.

Patterning, From Conifers to Consciousness: Turing's Theory and Order From Fluctuations.

This is a brief account of Turing's ideas on biological pattern and the events that led to their wider acceptance by biologists as a valid way to investigate developmental pattern, and of the value of theory more generally in biology. Periodic patterns have played a key role in this process, especially 2D arrays of oriented stripes, which proved a disappointment in theoretical terms in the case of Drosophila segmentation, but a boost to theory as applied to skin patterns in fish and model chemical reactions. The concept of "order from fluctuations" is a key component of Turing's theory, wherein pattern arises by selective amplification of spatial components concealed in the random disorder of molecular and/or cellular processes. For biological examples, a crucial point from an analytical standpoint is knowing the nature of the fluctuations, where the amplifier resides, and the timescale over which selective amplification occurs. The answer clarifies the difference between "inelegant" examples such as Drosophila segmentation, which is perhaps better understood as a programmatic assembly process, and "elegant" ones expressible in equations like Turing's: that the fluctuations and selection process occur predominantly in evolutionary time for the former, but in real time for the latter, and likewise for error suppression, which for Drosophila is historical, in being lodged firmly in past evolutionary events. The prospects for a further extension of Turing's ideas to the complexities of brain development and consciousness is discussed, where a case can be made that it could well be in neuroscience that his ideas find their most important application.

Development of the nervous system in hatchlings of Spadella cephaloptera (Chaetognatha), and implications for nervous system evolution in Bilateria.

Chaetognaths (arrow worms) play an important role as predators in planktonic food webs. Their phylogenetic position is unresolved, and among the numerous hypotheses, affinities to both protostomes and deuterostomes have been suggested. Many aspects of their life history, including ontogenesis, are poorly understood and, though some aspects of their embryonic and postembryonic development have been described, knowledge of early neural development is still limited. This study sets out to provide new insights into neurogenesis of newly hatched Spadella cephaloptera and their development during the following days, with attention to the two main nervous centers, the brain and the ventral nerve center. These were examined with immunohistological methods and confocal laser-scan microscopic analysis, using antibodies against tubulin, FMRFamide, and synapsin to trace the emergence of neuropils and the establishment of specific peptidergic subsystems. At hatching, the neuronal architecture of the ventral nerve center is already well established, whereas the brain and the associated vestibular ganglia are still rudimentary. The development of the brain proceeds rapidly over the next 6 days to a state that resembles the adult pattern. These data are discussed in relation to the larval life style and behaviors such as feeding. In addition, we compare the larval chaetognath nervous system and that of other bilaterian taxa in order to extract information with phylogenetic value. We conclude that larval neurogenesis in chaetognaths does not suggest an especially close relationship to either deuterostomes or protostomes, but instead displays many apomorphic features.

Consciousness as a Product of Evolution: Contents, Selector Circuits, and Trajectories in Experience Space.

Conscious experience can be treated as a complex unified whole, but to do so is problematic from an evolutionary perspective if, like other products of evolution, consciousness had simple beginnings, and achieved complexity only secondarily over an extended period of time as new categories of subjective experience were added and refined. The premise here is twofold, first that these simple beginnings can be investigated regardless of whether the ultimate source of subjective experience is known or understood, and second, that of the contents known to us, the most accessible for investigation will be those that are, or appear, most fundamental, in the sense that they resist further deconstruction or analysis. This would include qualia as they are usually defined, but excludes more complex experiences (here, formats) that are structured, or depend on algorithmic processes and/or memory. Vision and language for example, would by this definition be formats. More formally, qualia, but not formats, can be represented as points, lines, or curves on a topological experience space, and as domains in a configuration space representing a subset of neural correlates of consciousness, the selector circuits (SCs), responsible for ensuring that a particular experience is evoked rather than some other. It is a matter of conjecture how points in SC-space map to experience space, but both will exhibit divergence, insuring that a minimal distance separates points in experience space representing different qualia and the SCs that evoke them. An analysis of how SCs evolve over time is used to highlight the importance of understanding patterns of descent among putative qualia, i.e., their homology across species, and whether this implies descent from an ancestral experience, or ur-quale, that combines modes of experience that later came to be experienced separately. The analysis also provides insight into the function of consciousness as viewed from an evolutionary perspective, defined here in terms of the access it allows to regions of SC-space that would otherwise be unavailable to real brains, to produce consciously controlled behaviors that could otherwise not occur.

Evolving Consciousness: Insights From Turing, and the Shaping of Experience.

A number of conceptual difficulties arise when considering the evolutionary origin of consciousness from the pre-conscious condition. There are parallels here with biological pattern formation, where, according to Alan Turing's original formulation of the problem, the statistical properties of molecular-level processes serve as a source of incipient pattern. By analogy, the evolution of consciousness can be thought of as depending in part on a competition between alternative variants in the microstructure of synaptic networks and/or the activity patterns they generate, some of which then serve as neural correlates of consciousness (NCCs). Assuming that NCCs perform this function only if reliably ordered in a particular and precise way, Turing's formulation provides a useful conceptual framework for thinking about how this is achieved developmentally, and how changes in neural structure might correlate with change at the level of conscious experience. The analysis is largely silent concerning the nature and ultimate source of conscious experience, but shows that achieving sentience is sufficient to begin the process by which evolution elaborates and shapes that first experience. By implication, much of what evolved consciousness achieves in adaptive terms can in principle be investigated irrespective of whether or not the ultimate source of real-time experience is known or understood. This includes the important issue of how precisely NCCs must be structured to ensure that each evokes a particular experience as opposed to any other. Some terminological issues are clarified, including that of "noise," which here refers to the statistical variations in neural structure that arise during development, not to sensory noise as experienced in real time.

Basic features of the ancestral chordate brain: a protochordate perspective.

Basic features of the anterior nerve cord in amphioxus larvae are summarized to highlight its essential similarity with the vertebrate brain. Except for a pineal homolog, the amphioxus brain consists of a much simplified version of the ventral brainstem, including a region probably homologous with the hypothalamus, and a locomotory control center roughly comparable to the vertebrate tegmentum and reticulospinal system. Amphioxus has direct pathways for activating its locomotory circuits in response to mechanical stimuli via epithelial sensory cells, but this response is evidently modulated by inputs from diverse sensory-type cells located in the putative hypothalamic homolog, and from the lamellar body, the pineal homolog. This implies that a basic function of the amphioxus brain is to switch between locomotory activities, of which there are several, and the principal non-locomotory one, namely feeding. A similar involvement in switching between behavioral modes may thus have been a core brain function in ancestral chordates. Currently, however, incomplete knowledge of the physiology and behavior of amphioxus limits how effectively it can be used as an evolutionary model. Eye evolution is briefly discussed to illustrate how a better understanding of living forms can inform the evolutionary debate. An account of recent data on dorsoventral inversion is also included, as this bears directly on the question of where the chordate brain originated in relation to other structures. It now appears likely that key components of the ancestral brain were originally located around the mouth. A secondary repositioning of the latter would therefore have been required before a unitary brain could be assembled and internalized. This association between the mouth and the evolving brain reinforces the idea of a fundamental early connection between core brain structures and the control of feeding activity.

Amphioxus, motion detection, and the evolutionary origin of the vertebrate retinotectal map.

The axonal projection from the retina to the optic tectum maps visual information isomorphically from one to the other and serves as a model for the development of sensory maps more generally in the vertebrate brain. How or why this connection evolved is not known, nor why the midbrain is so important to the processing of visual information. Amphioxus is potentially informative here because its eye homolog, the frontal eye, also has a neural connection to a region of the brain now known to be homologous with the caudal diencephalon and midbrain. The frontal eye has only a one-dimensional receptor array, but simple alterations to the pattern and plane of cell division would have been sufficient to generate a structure more like the vertebrate retina. Accounting for the retinotectal map poses more of a problem. The hypothesis developed here is that this is best explained as a consequence of a prior association between the roof of the anterior nerve cord and an array of rhabdomeric photoreceptors, homologous with the Joseph cells of amphioxus, that were used by the common ancestor of amphioxus and vertebrates for detecting moving shadows. Hence, a rudimentary tectal map could have been present before the evolution of image-forming eyes and been coopted by them secondarily. Assuming the orientation of this map was fixed from the start relative to the external world, its retinal counterpart would have had to adjust to this to accommodate the image reversal that accompanies the conversion of a flat receptor array to a camera-type eye. Exploring this hypothesis further will require more information than is currently available on the Joseph cells, especially as to where and how their neural output is processed.

Interpreting amphioxus, and thoughts on ancestral chordate mouths and brains.

Amphioxus is increasingly important as a model for ancestral chordates. Nevertheless, it is secondarily modified in various ways, especially in the larva, whose small size has resulted in a rescaling and repositioning of structures. This is especially pronounced in the head region, where the mouth opens asymmetrically on the left side, leading to speculation that the mouth is secondarily derived, e.g. from a gill slit, and is hence not homologous with mouths in other animals. The available evidence does not, in the author's view, support this interpretation. A second issue is raised concerning the identity and function of the midbrain homolog, whose extent depends on whether greater weight is given to dorsal landmarks in the nerve cord or ventral ones. The presence of two sets of dorsal photoreceptors, the lamellar body and Joseph cells, functionally links the region they occupy to the vertebrate midbrain. The midbrain is currently suggested to be the brain region in which primary consciousness emerged during early vertebrate evolution, so the origin of its constituent cells is of special interest. Possible amphioxus homologs include the anterior-most group of dorsal bipolar cells (ADBs), which are apico-basally inverted (i.e. synapse-bearing neurites arise from the apical cell compartment) in the same fashion as cortical neurons in vertebrates. This may have been a crucial innovation for chordates, responsible for both improved sensory processing and, eventually, consciousness.

Locomotory control in amphioxus larvae: new insights from neurotransmitter data.

Amphioxus larvae have a midbrain-level locomotory control center whose overall organization is known from serial TEM reconstructions. How it functions has been a puzzle, owing to uncertainty as to the transmitters used by each class of neurons, but this has recently become clearer. We summarize what is now known, and correct past misconceptions: The large paired neurons at the core of the control center are glutamatergic, and hence excitatory, the commissural neurons are GABAergic, hence probably inhibitory, and both motoneurons and ipsilateral projection neurons are cholinergic, suggesting that the latter, a class of interneurons, may be derived evolutionarily from the former. The data clarify some aspects of how fast and slow swimming are controlled and prevented from interfering with one another, but leave open the source of pacemaker activity, which could reside in the large paired neurons or circuits associated with them. A unusual type of non-synaptic junction links the fast and slow systems, but how these junctions function is open to interpretation, depending chiefly on whether they act to couple adjacent cells independent of cell type, or can have differential effects that vary with cell type. Some evolutionary implications are discussed.

The origin of dopaminergic systems in chordate brains: insights from amphioxus.

The basic anatomy of the central nervous system (CNS) is well conserved within the vertebrates and differs in significant ways from that of non-vertebrate chordates. Of the latter, amphioxus is of special interest, being the best available stand-in for the basal chordate condition. Immunohistochemical and gene expression studies on the developing CNS of amphioxus embryos and larvae are now sufficiently advanced that we can begin to assign specific neurotransmitter phenotypes to neurons identified by transmission electron microscopy (TEM), and then compare the distribution of cell types to that in vertebrate brains. Here, by monitoring tyrosine hydroxylase (TH) transcripts and protein, along with serial TEM, we identify a population of catecholamine-containing neurons in the anterior nerve cord of amphioxus larvae and describe their pattern of synaptic inputs and outputs. Inputs parallel those to the large paired neurons that control the larval escape response, suggesting that the TH+ system functions as an accessory excitatory and perhaps modulatory pathway in larval locomotion, with the added feature of recruiting an assortment of additional interneurons to the circuitry. The TH+ cells probably contain either L-DOPA or dopamine, and correspond closely with a cell population known from previous work on adult amphioxus to be dopaminergic. This population lies in a CNS domain now thought to comprise a combined vertebrate diencephalon plus mesencephalon, the implication being that dopaminergic nuclei in both of these brain regions could derive from a single dien-mesencephalic population in the last common ancestor of amphioxus and vertebrates.