Navigating Central Oxytocin Transport: Known Realms and Uncharted Territories.

Complex mechanisms govern the transport and action of oxytocin (Oxt), a neuropeptide and hormone that mediates diverse physiologic processes. While Oxt exerts site-specific and rapid effects in the brain via axonal and somatodendritic release, volume transmission via CSF and the neurovascular interface can act as an additional mechanism to distribute Oxt signals across distant brain regions on a slower timescale. This review focuses on modes of Oxt transport and action in the CNS, with particular emphasis on the roles of perivascular spaces, the blood-brain barrier (BBB), and circumventricular organs in coordinating the triadic interaction among circulating blood, CSF, and parenchyma. Perivascular spaces, critical conduits for CSF flow, play a pivotal role in Oxt diffusion and distribution within the CNS and reciprocally undergo Oxt-mediated structural and functional reconstruction. While the BBB modulates the movement of Oxt between systemic and cerebral circulation in a majority of brain regions, circumventricular organs without a functional BBB can allow for diffusion, monitoring, and feedback regulation of bloodborne peripheral signals such as Oxt. Recognition of these additional transport mechanisms provides enhanced insight into the systemic propagation and regulation of Oxt activity.

Understanding electrical and chemical transmission in the brain.

The histochemical Falck-Hillarp method for the localization of dopamine (DA), noradrenaline (NA) and serotonin in the central nervous system (CNS) of rodents was introduced in the 1960s. It supported the existence of chemical neurotransmission in the CNS. The monoamine neurons in the lower brain stem formed monosynaptic ascending systems to the telencephalon and diencephalon and monoamine descending systems to the entire spinal cord. The monoamines were early on suggested to operate via synaptic chemical transmission in the CNS. This chemical transmission reduced the impact of electrical transmission. In 1969 and the 1970s indications were obtained that important modes of chemical monoamine communication in the CNS also took place through the extra-synaptic fluid, the extracellular fluid, and long-distance communication in the cerebrospinal fluid involving diffusion and flow of transmitters like DA, NA and serotonin. In 1986, this type of transmission was named volume transmission (VT) by Agnati and Fuxe and their colleagues, also characterized by transmitter varicosity and receptor mismatches. The short and long-distance VT pathways were characterized by volume fraction, tortuosity and clearance. Electrical transmission also exists in the mammalian CNS, but chemical transmission is in dominance. One electrical mode is represented by electrical synapses formed by gap junctions which represent low resistant passages between nerve cells. It allows for a more rapid passage of action potentials between nerve cells compared to chemical transmission. The second mode is based on the ability of synaptic currents to generate electrical fields to modulate chemical transmission. One aim is to understand how chemical transmission can be integrated with electrical transmission and how putative (aquaporin water channel, dopamine D2R and adenosine A2AR) complexes in astrocytes can significancy participate in the clearance of waste products from the glymphatic system. VT may also help accomplish the operation of the acupuncture meridians essential for Chinese medicine in view of the indicated existence of extracellular VT pathways.

Chemical cognition: chemoconnectomics and convergent evolution of integrative systems in animals.

Neurons underpin cognition in animals. However, the roots of animal cognition are elusive from both mechanistic and evolutionary standpoints. Two conceptual frameworks both highlight and promise to address these challenges. First, we discuss evidence that animal neural and other integrative systems evolved more than once (convergent evolution) within basal metazoan lineages, giving us unique experiments by Nature for future studies. The most remarkable examples are neural systems in ctenophores and neuroid-like systems in placozoans and sponges. Second, in addition to classical synaptic wiring, a chemical connectome mediated by hundreds of signal molecules operates in tandem with neurons and is the most information-rich source of emerging properties and adaptability. The major gap-dynamic, multifunctional chemical micro-environments in nervous systems-is not understood well. Thus, novel tools and information are needed to establish mechanistic links between orchestrated, yet cell-specific, volume transmission and behaviors. Uniting what we call chemoconnectomics and analyses of the cellular bases of behavior in basal metazoan lineages arguably would form the foundation for deciphering the origins and early evolution of elementary cognition and intelligence.

Wiring and Volume Transmission: An Overview of the Dual Modality for Serotonin Neurotransmission.

Serotonin is a neurotransmitter involved in the modulation of a multitude of physiological and behavioral processes. In spite of the relatively reduced number of serotonin-producing neurons present in the mammalian CNS, a complex long-range projection system provides profuse innervation to the whole brain. Heterogeneity of serotonin receptors, grouped in seven families, and their spatiotemporal expression pattern account for its widespread impact. Although neuronal communication occurs primarily at tiny gaps called synapses, wiring transmission, another mechanism based on extrasynaptic diffusion of neuroactive molecules and referred to as volume transmission, has been described. While wiring transmission is a rapid and specific one-to-one modality of communication, volume transmission is a broader and slower mode in which a single element can simultaneously act on several different targets in a one-to-many mode. Some experimental evidence regarding ultrastructural features, extrasynaptic localization of receptors and transporters, and serotonin-glia interactions collected over the past four decades supports the existence of a serotonergic system of a dual modality of neurotransmission, in which wiring and volume transmission coexist. To date, in spite of the radical difference in the two modalities, limited information is available on the way they are coordinated to mediate the specific activities in which serotonin participates. Understanding how wiring and volume transmission modalities contribute to serotonergic neurotransmission is of utmost relevance for the comprehension of serotonin functions in both physiological and pathological conditions.

Dynamical questions in volume transmission.

In volume transmission (or neuromodulation) neurons do not make one-to-one connections to other neurons, but instead simply release neurotransmitter into the extracellular space from numerous varicosities. Many well-known neurotransmitters including serotonin (5HT), dopamine (DA), histamine (HA), Gamma-Aminobutyric Acid (GABA) and acetylcholine (ACh) participate in volume transmission. Typically, the cell bodies are in one volume and the axons project to a distant volume in the brain releasing the neurotransmitter there. We introduce volume transmission and describe mathematically two natural homeostatic mechanisms. In some brain regions several neurotransmitters in the extracellular space affect each other's release. We investigate the dynamics created by this comodulation in two different cases: serotonin and histamine; and the comodulation of 4 neurotransmitters in the striatum and we compare to experimental data. This kind of comodulation poses new dynamical questions as well as the question of how these biochemical networks influence the electrophysiological networks in the brain.

Holographic Recording of Unslanted Volume Transmission Gratings in Acrylamide/Propargyl Acrylate Hydrogel Layers: Towards Nucleic Acids Biosensing.

The role of volume hydrogel holographic gratings as optical transducers in sensor devices for point-of-care applications is increasing due to their ability to be functionalized for achieving enhanced selectivity. The first step in the development of these transducers is the optimization of the holographic recording process. The optimization aims at achieving gratings with reproducible diffraction efficiency, which remains stable after reiterative washings, typically required when working with analytes of a biological nature or several step tests. The recording process of volume phase transmission gratings within Acrylamide/Propargyl Acrylate hydrogel layers reported in this work was successfully performed, and the obtained diffraction gratings were optically characterized. Unslanted volume transmission gratings were recorded in the hydrogel layers diffraction efficiencies; up to 80% were achieved. Additionally, the recorded gratings demonstrated stability in water after multiple washing steps. The hydrogels, after functionalization with oligonucleotide probes, yields a specific hybridization response, recognizing the complementary strand as demonstrated by fluorescence. Analyte-sensitive hydrogel layers with holographic structures are a promising candidate for the next generation of in vitro diagnostic tests.

Vasculature of the Suprachiasmatic Nucleus: Pathways for Diffusible Output Signals.

Transplant studies demonstrate unequivocally that the suprachiasmatic nucleus (SCN) produces diffusible signals that can sustain circadian locomotor rhythms. There is a vascular portal pathway between the SCN and the organum vasculosum of the lamina terminalis in mouse brain. Portal pathways enable low concentrations of neurosecretions to reach specialized local targets without dilution in the systemic circulation. To explore the SCN vasculature and the capillary vessels whereby SCN neurosecretions might reach portal vessels, we investigated the blood vessels (BVs) of the core and shell SCN. The arterial supply of the SCN differs among animals, and in some animals, there are differences between the 2 sides. The rostral SCN is supplied by branches from either the superior hypophyseal artery (SHpA) or the anterior cerebral artery or the anterior communicating artery. The caudal SCN is consistently supplied by the SHpA. The rostral SCN is drained by the preoptic vein, while the caudal is drained by the basal vein, with variations in laterality of draining vessels. In addition, several key features of the core and shell SCN regions differ: Median BV diameter is significantly smaller in the shell than the core based on confocal image measurements, and a similar trend occurs in iDISCO-cleared tissue. In the cleared tissue, whole BV length density and surface area density are significantly greater in the shell than the core. Finally, capillary length density is also greater in the shell than the core. The results suggest three hypotheses: First, the distinct arterial and venous systems of the rostral and caudal SCN may contribute to the in vivo variations of metabolic and neural activities observed in SCN networks. Second, the dense capillaries of the SCN shell are well positioned to transport blood-borne signals. Finally, variations in SCN vascular supply and drainage may contribute to inter-animal differences.

Neuropeptidergic control circuits in the spinal cord for male sexual behaviour: Oxytocin-gastrin-releasing peptide systems.

The neuropeptidergic mechanisms controlling socio-sexual behaviours consist of complex neuronal circuitry systems in widely distributed areas of the brain and spinal cord. At the organismal level, it is now becoming clear that "hormonal regulations" play an important role, in addition to the activation of neuronal circuits. The gastrin-releasing peptide (GRP) system in the lumbosacral spinal cord is an important component of the neural circuits that control penile reflexes in rats, circuits that are commonly referred to as the "spinal ejaculation generator (SEG)." Oxytocin, long known as a neurohypophyseal hormone, is now known to be involved in the regulation of socio-sexual behaviors in mammals, ranging from social bonding to empathy. However, the functional interaction between the SEG neurons and the hypothalamo-spinal oxytocin system remains unclear. Oxytocin is known to be synthesised mainly in hypothalamic neurons and released from the posterior pituitary into the circulation. Oxytocin is also released from the dendrites of the neurons into the hypothalamus where they have important roles in social behaviours via non-synaptic volume transmission. Because the most familiar functions of oxytocin are to regulate female reproductive functions including parturition, milk ejection, and maternal behaviour, oxytocin is often thought of as a "feminine" hormone. However, there is evidence that a group of parvocellular oxytocin neurons project to the lower spinal cord and control male sexual function in rats. In this report, we review the functional interaction between the SEG neurons and the hypothalamo-spinal oxytocin system and effects of these neuropeptides on male sexual behaviour. Furthermore, we discuss the finding of a recently identified, localised "volume transmission" role of oxytocin in the spinal cord. Findings from our studies suggest that the newly discovered "oxytocin-mediated spinal control of male sexual function" may be useful in the treatment of erectile and ejaculatory dysfunction.

Brain Structure and Function: Insights from Chemical Neuroanatomy.

We present a brief historical and epistemological outline of investigations on the brain's structure and functions. These investigations have mainly been based on the intermingling of chemical anatomy, new techniques in the field of microscopy and computer-assisted morphometric methods. This intermingling has enabled extraordinary investigations to be carried out on brain circuits, leading to the development of a new discipline: "brain connectomics". This new approach has led to the characterization of the brain's structure and function in physiological and pathological conditions, and to the development of new therapeutic strategies. In this context, the conceptual model of the brain as a hyper-network with a hierarchical, nested architecture, arranged in a "Russian doll" pattern, has been proposed. Our investigations focused on the main characteristics of the modes of communication between nodes at the various miniaturization levels, in order to describe the brain's integrative actions. Special attention was paid to the nano-level, i.e., to the allosteric interactions among G protein-coupled receptors organized in receptor mosaics, as a promising field in which to obtain a new view of synaptic plasticity and to develop new, more selective drugs. The brain's multi-level organization and the multi-faceted aspects of communication modes point to an emerging picture of the brain as a very peculiar system, in which continuous self-organization and remodeling take place under the action of external stimuli from the environment, from peripheral organs and from ongoing integrative actions.

Volume-transmitted GABA waves pace epileptiform rhythms in the hippocampal network.

Mechanisms that entrain and pace rhythmic epileptiform discharges remain debated. Traditionally, the quest to understand them has focused on interneuronal networks driven by synaptic GABAergic connections. However, synchronized interneuronal discharges could also trigger the transient elevations of extracellular GABA across the tissue volume, thus raising tonic conductance (Gtonic) of synaptic and extrasynaptic GABA receptors in multiple cells. Here, we monitor extracellular GABA in hippocampal slices using patch-clamp GABA "sniffer" and a novel optical GABA sensor, showing that periodic epileptiform discharges are preceded by transient, region-wide waves of extracellular GABA. Neural network simulations that incorporate volume-transmitted GABA signals point to a cycle of GABA-driven network inhibition and disinhibition underpinning this relationship. We test and validate this hypothesis using simultaneous patch-clamp recordings from multiple neurons and selective optogenetic stimulation of fast-spiking interneurons. Critically, reducing GABA uptake in order to decelerate extracellular GABA fluctuations-without affecting synaptic GABAergic transmission or resting GABA levels-slows down rhythmic activity. Our findings thus unveil a key role of extrasynaptic, volume-transmitted GABA in pacing regenerative rhythmic activity in brain networks.

Nanoscopic dopamine transporter distribution and conformation are inversely regulated by excitatory drive and D2 autoreceptor activity.

The nanoscopic organization and regulation of individual molecular components in presynaptic varicosities of neurons releasing modulatory volume neurotransmitters like dopamine (DA) remain largely elusive. Here we show, by application of several super-resolution microscopy techniques to cultured neurons and mouse striatal slices, that the DA transporter (DAT), a key protein in varicosities of dopaminergic neurons, exists in the membrane in dynamic equilibrium between an inward-facing nanodomain-localized and outward-facing unclustered configuration. The balance between these configurations is inversely regulated by excitatory drive and DA D2 autoreceptor activation in a manner dependent on Ca2+ influx via N-type voltage-gated Ca2+ channels. The DAT nanodomains contain tens of transporters molecules and overlap with nanodomains of PIP2 (phosphatidylinositol-4,5-bisphosphate) but show little overlap with D2 autoreceptor, syntaxin-1, and clathrin nanodomains. The data reveal a mechanism for rapid alterations of nanoscopic DAT distribution and show a striking link of this to the conformational state of the transporter.

A Multidisciplinary Hypothesis about Serotonergic Psychedelics. Is it Possible that a Portion of Brain Serotonin Comes From the Gut?

Here we present a complex hypothesis about the psychosomatic mechanism of serotonergic psychedelics. Serotonergic psychedelics affect gut microbes that produce a temporary increase of 5-HT by their host enterochromaffin cells (ECs). This increased 5-HT production-which is taken up and distributed by platelets-may work as a hormone-like regulatory signal that could influence membrane permeability in the host organs and tissues and in the brain. Increased plasma 5-HT levels could enhance permeability of the blood-brain barrier (BBB). Transiently increased permeability of the BBB allows for plasma 5-HT to enter the central nervous system (CNS) and be distributed by the volume transmission. Next, this gut-derived 5-HT could modulate excitatory and inhibitory neurotransmission and produce special network disintegration in the CNS. This transient perturbation of the normal neural hierarchy allows patients access to suppressed fear information and perform an emotional reset, in which the amygdale may have a key role.

A Magnetic Nanoparticle-Doped Photopolymer for Holographic Recording.

Functionalised holograms are important for applications utilising smart diffractive optical elements for light redirection, shaping and in the development of sensors/indicators. This paper reports on holographic recording in novel magnetic nanocomposites and the observed temperature change in dry layers and liquid samples exposed to alternating magnetic field (AMF). The nanocomposite consists of N-isopropylacrylamide (NIPA)-based polymer doped with magnetic nanoparticles (MNPs), and local heating is achieved through magnetic induction. Here, volume transmission holographic gratings (VTHGs) are recorded with up to 24% diffraction efficiency (DE) in the dry layers of magnetic nanocomposites. The dry layers and liquid samples are then exposed to AMF. Efficient heating was observed in the liquid samples doped with Fe3O4 MNPs of 20 nm average size where the temperature increased from 27 °C to 64 °C after 300 s exposure to 111 mT AMF. The temperature increase in the dry layers doped with the same nanoparticles after exposure to 4.4 mT AMF was observed to be 6 °C. No temperature change was observed in the undoped layers. Additionally, we have successfully recorded Denisyuk holograms in the magnetic nanocomposite materials. The results reveal that the magnetic nanocomposite layers are suitable for recording holograms and need further optimisation in developing holographic indicators for mapping AMFs.

The Nature of Noradrenergic Volume Transmission From Locus Coeruleus to Brainstem Mesencephalic Trigeminal Sensory Neurons.

Noradrenergic neurons in the locus coeruleus (LC) release noradrenaline (NA) that acts via volume transmission to activate extrasynaptic G-protein coupled receptors (GPCRs) in target cells throughout the brain. As the closest projection, the dorsal LC laterally adjoins the mesencephalic trigeminal nucleus (MTN), in which proprioceptive primary sensory neurons innervating muscle spindles of jaw-closing muscles are exceptionally located. MTN neurons express α2-adrenergic receptors (α2-ARs) and display hyperpolarization-activated cyclic nucleotide-gated (HCN) currents (Ihs), which is downregulated by α2-AR activation. To quantify the activity-dependent outcome of volume transmission of NA from LC to MTN, we investigated how direct LC activation inhibits Ih in MTN neurons by performing dual whole-cell recordings from LC and MTN neurons. Repetition of 20 Hz spike-train evoked with 1-s current-pulse in LC neurons every 30 s resulted in a gradual decrease in Ih evoked every 30 s, revealing a Hill-type relationship between the number of spike-trains in LC neurons and the degree of Ih inhibition in MTN neurons. On the other hand, when microstimulation was applied in LC every 30 s, an LC neuron repeatedly displayed a transient higher-frequency firing followed by a tonic firing at 5-10 Hz for 30 s. This subsequently caused a similar Hill-type inhibition of Ih in the simultaneously recorded MTN neuron, but with a smaller Hill coefficient, suggesting a lower signal transduction efficacy. In contrast, 20 Hz activity induced by a 1-s pulse applied every 5-10 s caused only a transient facilitation of Ih inhibition followed by a forced termination of Ih inhibition. Thus, the three modes of LC activities modulated the volume transmission to activate α2-adrenergic GPCR to differentially inhibit Ih in MTN neurons.

Intercellular Communication in the Central Nervous System as Deduced by Chemical Neuroanatomy and Quantitative Analysis of Images: Impact on Neuropharmacology.

In the last decades, new evidence on brain structure and function has been acquired by morphological investigations based on synergic interactions between biochemical anatomy approaches, new techniques in microscopy and brain imaging, and quantitative analysis of the obtained images. This effort produced an expanded view on brain architecture, illustrating the central nervous system as a huge network of cells and regions in which intercellular communication processes, involving not only neurons but also other cell populations, virtually determine all aspects of the integrative function performed by the system. The main features of these processes are described. They include the two basic modes of intercellular communication identified (i.e., wiring and volume transmission) and mechanisms modulating the intercellular signaling, such as cotransmission and allosteric receptor-receptor interactions. These features may also open new possibilities for the development of novel pharmacological approaches to address central nervous system diseases. This aspect, with a potential major impact on molecular medicine, will be also briefly discussed.

Striatal Cholinergic Signaling in Time and Space.

The cholinergic interneurons of the striatum account for a small fraction of all striatal cell types but due to their extensive axonal arborization give the striatum the highest content of acetylcholine of almost any nucleus in the brain. The prevailing theory of striatal cholinergic interneuron signaling is that the numerous varicosities on the axon produce an extrasynaptic, volume-transmitted signal rather than mediating rapid point-to-point synaptic transmission. We review the evidence for this theory and use a mathematical model to integrate the measurements reported in the literature, from which we estimate the temporospatial distribution of acetylcholine after release from a synaptic vesicle and from multiple vesicles during tonic firing and pauses. Our calculations, together with recent data from genetically encoded sensors, indicate that the temporospatial distribution of acetylcholine is both short-range and short-lived, and dominated by diffusion. These considerations suggest that acetylcholine signaling by cholinergic interneurons is consistent with point-to-point transmission within a steep concentration gradient, marked by transient peaks of acetylcholine concentration adjacent to release sites, with potential for faithful transmission of spike timing, both bursts and pauses, to the postsynaptic cell. Release from multiple sites at greater distance contributes to the ambient concentration without interference with the short-range signaling. We indicate several missing pieces of evidence that are needed for a better understanding of the nature of synaptic transmission by the cholinergic interneurons of the striatum.

Neurobiological reduction: From cellular explanations of behavior to interventions.

Scientific reductionism, the view that higher level functions can be explained by properties at some lower-level or levels, has been an assumption of nervous system analyses since the acceptance of the neuron doctrine in the late 19th century, and became a dominant experimental approach with the development of intracellular recording techniques in the mid-20th century. Subsequent refinements of electrophysiological approaches and the continual development of molecular and genetic techniques have promoted a focus on molecular and cellular mechanisms in experimental analyses and explanations of sensory, motor, and cognitive functions. Reductionist assumptions have also influenced our views of the etiology and treatment of psychopathologies, and have more recently led to claims that we can, or even should, pharmacologically enhance the normal brain. Reductionism remains an area of active debate in the philosophy of science. In neuroscience and psychology, the debate typically focuses on the mind-brain question and the mechanisms of cognition, and how or if they can be explained in neurobiological terms. However, these debates are affected by the complexity of the phenomena being considered and the difficulty of obtaining the necessary neurobiological detail. We can instead ask whether features identified in neurobiological analyses of simpler aspects in simpler nervous systems support current molecular and cellular approaches to explaining systems or behaviors. While my view is that they do not, this does not invite the opposing view prevalent in dichotomous thinking that molecular and cellular detail is irrelevant and we should focus on computations or representations. We instead need to consider how to address the long-standing dilemma of how a nervous system that ostensibly functions through discrete cell to cell communication can generate population effects across multiple spatial and temporal scales to generate behavior.

Assessment of astrocytes as a mediator of memory and learning in rodents.

The classical view of astrocytes is that they provide supportive functions for neurons, transporting metabolites and maintaining the homeostasis of the extracellular milieu. This view is gradually changing with the advent of molecular genetics and optical methods allowing interrogation of selected cell types in live experimental animals. An emerging view that astrocytes additionally act as a mediator of synaptic plasticity and contribute to learning processes has gained in vitro and in vivo experimental support. Here we focus on the literature published in the past two decades to review the roles of astrocytes in brain plasticity in rodents, whereby the roles of neurotransmitters and neuromodulators are considered to be comparable to those in humans. We outline established inputs and outputs of astrocytes and discuss how manipulations of astrocytes have impacted the behavior in various learning paradigms. Multiple studies suggest that the contribution of astrocytes has a considerably longer time course than neuronal activation, indicating metabolic roles of astrocytes. We advocate that exploring upstream and downstream mechanisms of astrocytic activation will further provide insight into brain plasticity and memory/learning impairment.

Expression Patterns of the Neuropeptide Urocortin 3 and Its Receptor CRFR2 in the Mouse Central Auditory System.

Sensory systems have to be malleable to context-dependent modulations occurring over different time scales, in order to serve their evolutionary function of informing about the external world while also eliciting survival-promoting behaviors. Stress is a major context-dependent signal that can have fast and delayed effects on sensory systems, especially on the auditory system. Urocortin 3 (UCN3) is a member of the corticotropin-releasing factor family. As a neuropeptide, UCN3 regulates synaptic activity much faster than the classic steroid hormones of the hypothalamic-pituitary-adrenal axis. Moreover, due to the lack of synaptic re-uptake mechanisms, UCN3 can have more long-lasting and far-reaching effects. To date, a modest number of studies have reported the presence of UCN3 or its receptor CRFR2 in the auditory system, particularly in the cochlea and the superior olivary complex, and have highlighted the importance of this stress neuropeptide for protecting auditory function. However, a comprehensive map of all neurons synthesizing UCN3 or CRFR2 within the auditory pathway is lacking. Here, we utilize two reporter mouse lines to elucidate the expression patterns of UCN3 and CRFR2 in the auditory system. Additional immunolabelling enables further characterization of the neurons that synthesize UCN3 or CRFR2. Surprisingly, our results indicate that within the auditory system, UCN3 is expressed predominantly in principal cells, whereas CRFR2 expression is strongest in non-principal, presumably multisensory, cell types. Based on the presence or absence of overlap between UCN3 and CRFR2 labeling, our data suggest unusual modes of neuromodulation by UCN3, involving volume transmission and autocrine signaling.