Sturgeons are an ancient and basal group of ray-finned fishes whose morphology is critical to our understanding of the evolution of teleosts, which form most of the ray-finned fish radiation. In sturgeons, the pallium, or area dorsalis, consists of lateral, medial, and posterior divisions. The connections of these pallial divisions were determined by injections of biotinylated dextran amine and DiI. All three divisions receive olfactory input, with the posterior division receiving the bulk of the secondary olfactory projections. All three pallial divisions also have extensive intratelencephalic connections, as well as input from the thalamus and the posterior tubercle. The posterior division also receives sparse input from the midbrain and secondary gustatory nucleus of the isthmus. All three pallial divisions project heavily to the hypothalamus of the ipsilateral inferior lobe, and more sparsely to the ipsilateral optic tectum, posterior tubercle, and midbrain tegmentum. Cytological and connectional data provide no evidence that either a dorsal or central pallial division exists in sturgeons, and these pallial divisions likely arose in ancestral teleosts.

Two lateral mouse brain sagittal sections labeled, respectively, for Lypd1 (A) and Kcnab3 (B). (A) Lypd1 labels positively cortical Layers 2 and 5 (less heavily), but only at limbic mesocortical sites, forming the limbic ring that separates the unlabeled isocortex (IsoCx) from the unlabeled hippocampal and olfactory allocortex (Hi, Sub, ERh; OlfCx). The limbic ring is marked twice as the insula (Ins, MCx) and the postrhinal area (PoRh, MCx). (B) Kcnab3, contrarily, labels selectively various layers of the isocortex (IsoCx) and the hippocampal allocortex (Hi, Sub, ERh) but leaves unlabeled selectively the insula (Ins, MCx) and postrhinal (PoRh, MCx) areas of the limbic mesocortical ring, being thus a negative marker for this sort of cortex.

Alvinocaridid shrimps colonize hydrothermal vent fields along the Mid-Atlantic Ridge (MAR), where they experience extreme environmental conditions including high hydrostatic pressure, steep thermal and chemical gradients, and dim light. While various aspects of the biology of the dominant vent shrimp Rimicaris exoculata, such as general anatomy, microhabitat, symbiotic relationships, or trophic networks, have been extensively studied, little is known about how brain organization varies among alvinocaridid species occupying different ecological niches. Here, we investigated whether vent shrimps share common neuroanatomical features and whether differences in brain and vascular organization reflect ecological diversification among species. Using histology, immunohistochemistry, µCT scans, and 3D reconstruction, we compared the brain and the neurovascular system (cor frontale) in four MAR species with distinct ecologies (R. exoculata, Rimicaris chacei, Mirocaris fortunata, Alvinocaris markensis) and in the coastal relative Palaemon elegans. Our results reveal strong commonalities in brain organization among vent shrimps. All MAR species display a compact brain architecture with hypertrophied mushroom bodies (MBs), suggesting enhanced multisensory integration and memory capacities. Corresponding to the low-light environment, the visual neuropils (lamina, medulla, lobula) are smaller than those of the coastal species P. elegans. In all species living near hydrothermal vents, the retina exhibits modifications; it extends along the dorsal part of the cephalothorax, and its length varies according to the species, being the longest in the genus Rimicaris. The lobula satellite neuropil is absent in all alvinocaridids, suggesting a limited capacity to track moving visual signals. The olfactory lobes exhibit a common glomerular organization but differ among species in quantitative parameters, such as volume and number of glomeruli. The neurovascular system shows marked interspecific variation; the cor frontale differs in size and volume among hydrothermal vent species but retains a conserved subdivision of central arteries surrounding a longitudinal muscle bundle. We conclude that the observed variations in the volume of neuropils and arrangement of the vascular system likely reflect the ecological divergence related to microhabitat conditions and trophic strategies near hydrothermal emissions.

Mechanosensory innervation of the oral cavity enables us to detect the texture and location of the foods we consume. Our goal was to find a genetic identifier for a mechanoreceptive subtype. Toward this goal, we examined neurons expressing the neurotrophin receptor TrkC (TrkC-tdTomato), parvalbumin (Pvalb), or glutamate transporter, Vglut3. We found that the majority of Pvalb-lineage, Vglut3-lineage, and TrkC-tdTomato neurons innervating the tongue are separate trigeminal neuron populations. Because different papilla types contribute to food detection in different ways, we examined the innervation patterns of these neuron subtypes. We found that Pvalb-lineage and Vglut3-lineage neurons were fungiform papilla-specific, whereas TrkC+ fibers innervated both papilla types. We found that all Pvalb-lineage nerve fibers are labeled with neurofilament heavy chain (NFH), have axons that are surrounded by myelin basic protein, and lack calcitonin gene-related peptide. This indicates that these are all myelinated neurons. In contrast, Vglut3-lineage and TrkC-tdTomato nerve fibers were a mixed population of NFH+ nerve fibers and NFH- nerve fibers, as well as myelinated and unmyelinated nerve fibers. When we examined the axonal ending morphologies of the three genetic populations, we found that the Pvalb-lineage neurons consistently displayed a "basket-like" axonal ending that surrounded the taste bud. While TrkC+ and Vglut3-lineage neurons sometimes displayed this "basket-like" axonal ending, other axonal endings lacked this structure. When compared across groups, the Vglut3-lineage axonal endings displayed the greatest degree of morphological variability. Collectively, these data show that Pvalb-lineage may be a genetic identifier for a single myelinated mechanoreceptor innervating fungiform papillae.

Ceramide synthases (CerS) are key enzymes in sphingolipid metabolism that regulate fundamental cellular processes, including apoptosis, cell growth, and homeostasis. Among the six known mammalian isoforms (CerS1-CerS6), CerS5 has been particularly well studied for its involvement in the synthesis of the sphingolipid C16-ceramide. However, its expression, localization, and functional significance of CerS5 in the retina remain unclear. In the present study, we investigated the presence, distribution, and functional role of CerS5 in mouse retina using CerS5 knockout (KO) mice. We performed quantitative polymerase chain reaction, X-gal staining, and immunohistochemistry to analyze the expression and localization. Electroretinography (ERG) was employed to assess the impact of CerS5 deficiency on retinal function. Our results demonstrated that CerS5 is localized to the inner nuclear layer and ganglion cell layer, co-localizing with horizontal cells and specific subsets of amacrine and ganglion cells. The retina of CerS5 KO mice showed a reduction in overall thickness, with significant thinning observed in all retinal layers except the photoreceptor, whereas the outer plexiform layer showed increased thickness. Despite these structural alterations, ERG recordings revealed no significant changes in retinal function. These findings suggest that CerS5 contributes to the maintenance of retinal structural integrity, particularly through its presence in specific retinal cell types, whereas its loss does not markedly impair retinal function in adult mice. The observed structural alterations highlight its potential role in retinal physiology and possible implications for retinal pathophysiology, warranting further investigation into compensatory mechanisms by other ceramide synthase isoforms.

Dopamine signaling through dopamine 1 receptors (D1Rs) and dopamine 2 receptors (D2Rs) regulates hippocampal synaptic plasticity underlying learning and memory, yet their subcellular localization within the hippocampus is unknown. Here, we performed electron microscopic immunocytochemistry to elucidate the distribution of D1R and D2R in subregions of the mouse hippocampus. In CA1 and CA3 stratum radiatum (SR), D1R- and D2R-immunoreactivity was found primarily on pyramidal cell dendritic spines and unmyelinated axons, and to a lesser extent in axon terminals and glia. In both regions, D1R-labeled terminals formed predominantly asymmetric (excitatory-type) synapses on dendritic spines, whereas D2R-labeled terminals formed mainly symmetric (inhibitory-type) synapses on pyramidal cell dendritic shafts. In the dentate gyrus (DG) hilus, D1R labeling was almost exclusively found in unmyelinated axons and glia. D2R immunoreactivity in the hilus similarly was present in unmyelinated axons and glia but was also detected in dendritic spines originating from mossy cells and in terminals forming symmetric synapses. These findings indicate that dopamine receptors are positioned to influence excitatory and inhibitory signaling in the murine hippocampus. As D1R and D2R exert opposing effects on neuronal signaling, their localization on pyramidal neuron compartments provides a structural substrate for bidirectional modulation of synaptic plasticity and pyramidal cell activity. In addition, the presence of D2Rs on inhibitory terminals contacting pyramidal neurons and hilar interneurons suggests a role in regulating inhibitory circuitry within the hippocampus.

The neurotransmitters vasotocin, oxytocin, dopamine, and serotonin are widely involved in vertebrate social behavior, and changes in their abundance and distribution in the brain have been linked to the evolution of complex sociality. Reptiles provide an excellent system in which to investigate the neural mechanisms of social living. Using immunohistochemistry, we compare distributions of these transmitters in two skinks differing primarily in social ecology: the family-living Liopholis whitii and the solitary Eulpamrus quoyii. We describe patterns of immunopositive signal for both cell bodies and fibers across the entire brain (excluding the olfactory bulbs). In both species, vasotocin and oxytocin were found in the preoptic area, paraventricular nucleus, supraoptic nucleus, dorsomedial hypothalamus, and supraoptic decussation, as well as surrounding the lateral forebrain bundle. Tyrosine hydroxylase (a marker for dopamine) was found in the paraventricular organ nucleus, substantia nigra, and ventral tegmental area, and serotonin was found in the raphe nuclei and superior reticular field. We found novel oxytocin cell groups in the dorsomedial hypothalamus and cerebellum of L. whitii, and novel serotonin signal in the red nucleus of E. quoyii. Immunopositive signals found only in L. whitii also include vasotocin in the ventral tegmental area, tyrosine hydroxylase in the interpeduncular nucleus, and serotonin in the suprachiasmatic nucleus. The qualitatively greater abundance of these transmitters in the family-living L. whitii suggests that these molecules may have played an important role in the evolution of social behavior in these skinks and provides a foundation for broader comparisons across the social skinks.

Mosquitoes are major disease vectors that pose significant threats to human health. Recent efforts to understand the neurobiology of these insects have utilized modern research tools to advance our knowledge of the neural circuits underlying ecologically relevant behaviors in adults. However, the neurobiology of mosquito larvae remains relatively unexplored. This report focuses on the neuromuscular system of mosquito larvae, providing a morphological analysis of the abdominal neuromuscular system in Aedes aegypti. We identified 24 muscle fibers per hemi-segment and detailed their anchoring points and orientations. Using immunocytochemistry, the organization and innervation of abdominal body-wall muscles were reconstructed. Two primary nerve branches emerged from the ventral nerve cord in each segment. The branches, trajectories, and targets of the two primary nerves were named and described. At the synaptic level, motor axons arborization on muscle fibers were quantitatively analyzed. Motor axons branched extensively on muscle fibers, covering up to 90% of large fibers' length and approximately 30% of small fibers. Varicosities along axonal branches showed considerable size variation, with antibody labeled active zones present in all varicosities. Furthermore, active zone densities per varicosity on smaller muscle fibers were significantly higher than those on larger fibers. These observations provide a functional and morphological framework for understanding the physiology of mosquito larval locomotion.

Habituation is a simple form of nonassociative learning that is characterized by a decrease in response to a repetitive stimulus. As escape responses can be energetically costly and disruptive to normal behavior, it is important that prospective prey learn whether a perceived stimulus is a genuine threat or an innocuous stimulus that they can ignore. In response to a visual looming stimulus, larval zebrafish perform a characteristic escape swim that reliably habituates, and because they are small and transparent, they have been an important model for characterizing brain-wide activity patterns during habituation. In this study, we explore the spatial properties of visual adaptation to gauge whether it is mediated by local, regional, or brain-wide circuits. We present repetitive visual loom stimuli either in a fixed position in visual space or in variable positions, while also performing brain-wide calcium imaging. Across the brain, we identify both neural responses that are specific to looms at particular positions within the visual field and responses that occur regardless of where the loom is presented. By quantifying the degree of adaptation across these responses, we show that brain-wide adaptation occurs more rapidly when the position of the loom remains unchanged and that alternate looms occurring in different parts of the visual field minimally contribute to adaptation for looms at the original position. We found that the tectum, homologous to the superior colliculus, has response profiles and spatial sensitivity indicative of important contributions to this position-specific visual adaptation.

The endocranial anatomy of oviraptorosaurian theropods, and their inferred sensory adaptations, has played a key role in understanding the origin of bird-like senses among avian ancestors. However, within oviraptorosaurs, sampling has thus far been limited, and a major gap exists in our understanding of the endocranial anatomy of an enigmatic subclade, Caenagnathidae. Here, we describe the inner ear morphology of three caenagnathid oviraptorosaurs, spanning a wide range of stratigraphy and body size. Our data show that the incipiently bird-like inner ears of caenagnathids were relatively similar both over the course of >10 million years and over multiple orders of magnitude of body mass. Overall, the caenagnathids sampled here show similarities to other maniraptoran theropods and particularly other oviraptorosaurs, but with some key differences that possibly speak to behavioral differences in these groups. In particular, a reduced cochlear duct in the most complete labyrinth suggests restricted hearing sensitivity in caenagnathids, which is at odds with previous suggestions that they were more predatory than oviraptorids. Our results highlight the independent acquisition of some bird-like features along the avian stem and show that late-diverging members of these clades may exhibit divergent features that do not characterize the diversity within the entire group.

The primate anterior entorhinal cortex (EC) receives rich projections from the amygdala and from multimodal association areas, including the medial prefrontal, anterior cingulate, and orbitofrontal cortices. Axon terminations from these structures on the anterior EC facilitate processing of the emotional aspects of stimuli and events. The EC projects to hippocampus, which is associated with episodic memory. Processing in the anterior EC is modulated by inhibitory neurons, which in primates express the calcium-binding proteins (CBPs): calretinin (CR), or calbindin (CB) or parvalbumin (PV), which collectively account for most inhibitory neurons in the primate cortex. Here, stereological analysis of these neurochemical classes of inhibitory neurons in the anterior half of EC in humans revealed similar patterns as in rhesus monkeys. In both primate species, the densest neuronal subpopulation of presumed inhibitory neurons expressed CR, followed by CB, and lastly by PV. In both species CR neurons were most prevalent in layers I and II, CB neurons in layers II and III and PV neurons in the middle-deep layers. Moreover, the medial and lateral sectors of the anterior EC had different densities of neurons expressing these CBPs. Further analysis revealed that in the human anterior EC, virtually all PV neurons expressed the GABAergic marker GAD67/GAD1 (glutamate decarboxylase 67/glutamate decarboxylase 1), whereas only two-thirds of CB neurons and only one-third of CR neurons colocalized with GAD67/GAD1. In the entire neuronal population of the anterior half of human EC estimated by stereology, 10% expressed GAD67/GAD1, comparable to the collective population of CBP-positive neurons that colocalized with GAD67/GAD1. These findings reveal that the medial and lateral sectors of anterior EC have distinct inhibitory microenvironments, which likely affect the processing of input and output of hippocampus.

Octopamine (OA), a biogenic amine functionally similar to vertebrate norepinephrine, plays an important role in invertebrate neurophysiology. Previous reports of putative octopaminergic cells in the pond snail, Lymnaea stagnalis, have revealed inconsistencies, prompting our investigation that combined in situ hybridization chain reaction with traditional immunohistochemical methods. We mapped tyramine β-hydroxylase (TBH) mRNA, the corresponding TBH protein, and the OA neurotransmitter. Approximately 40 neurons were labeled by all three methods and are presumably genuinely octopaminergic, including several neurons with previous electrophysiological evidence that OA is their neurotransmitter. Our results also revealed approximately another 40 cells that only showed evidence of mRNA and enzyme labeling, but no neurotransmitter, and another five to eight cells that only labeled for OA. Some of these results are likely explained by antibody cross-reactivity, but multiple TBH isoforms (some of which may not produce OA) or regulatory mechanisms that block TBH function also need to be considered. Overall, this study underlines the need for a more nuanced interpretation of neuroanatomical mapping and provides new insights into the organization of molluscan octopaminergic systems.

To navigate its environment, an animal extracts salient information from sounds using temporal and intensity cues. In birds, the nucleus laminaris (NL) detects the submillisecond differences in the arrival time of sound to the two ears, the interaural time differences (ITDs), to localize sounds. This ability is facilitated by inhibitory long-range projection neurons from the ipsilateral superior olivary nucleus (SON) that enable NL neurons to remain sensitive to ITDs across a large range of sound intensities. It is well known that the excitatory inputs to NL, from nucleus magnocellularis (NM), innervate a narrow isofrequency band along the ITD axis. However, the organization of the inhibitory input from the SON remains largely unknown. We analyzed the innervation pattern of individual axons from SON neurons within the chicken NL. SON axonal arborizations vary greatly in size and topographic organization. On average, an inhibitory SON neuron innervates one-third of both the tonotopic and ITD axes, markedly larger target regions than do the excitatory inputs from NM. Unlike the excitatory axons that are confined to one dendritic lamina (separating inputs from the two ears), most SON cells innervate both laminae to similar extents, as well as the somata of the NL neurons. In addition, we found that some NL-projecting SON neurons also send collateral axons to NM or the nucleus angularis. The pattern of synapses along SON axons suggests that the inhibitory activity of individual NL neurons is shaped by many SON neurons. A single SON neuron contributes only a small proportion of the inhibition on each NL neuron. This broad innervation pattern of SON neurons is well-suited to control the overall activity of NL, supporting accurate ITD detection in a broad range of sound environments.

The habenula (Hb) has high-intensity mu opioid binding and receptor (MOR) expression. It contains medial and lateral subdivisions (MHb and LHb, respectively), yet the details of MOR localization across these regions remain debated. MHb and LHb participate in largely non-overlapping neural circuits; therefore, accurately resolving MOR expression across them is critical for understanding how MOR ligands impact behaviors. Here we utilized in situ hybridization (ISH) and immunocytochemistry (ICC) to systematically map Oprm1 mRNA and MOR protein throughout the habenular complex. We studied rat and mouse tissue to evaluate expression across two common research species. We also performed parallel mapping in Oprm1-Venus/Venus mice. Importantly, we found mRNA and protein in both MHb and LHb in both species. In rat, 39 ± 3% and 21 ± 4% of cells expressed Oprm1 in MHb and LHb, respectively. These proportions were greater in mouse: 57 ± 1% (MHb) and 32 ± 4% (LHb). Oprm1 puncta per positive cell were greater in MHb compared to LHb for both rat and mouse (p < 0.0001). The highest intensity labeling was localized along the lateral edge of the MHb for all methods. ICC showed MOR localized to fibers and somata in both regions. In LHb, MOR labeling was most dense in intermediate sections along the anterior-posterior (AP) axis. In rats we also observed greater labeling in dorsal LHb at intermediate AP levels and medial LHb more posteriorly. These results indicate that both MHb and LHb can contribute to MOR-mediated actions through their respective circuits.

Proline-rich transmembrane protein 2 (PRRT2) plays a pivotal role in the control of voluntary movements, as PRRT2 mutations cause paroxysmal kinesigenic dyskinesia (PKD) in a loss-of-function manner. Although the cerebellum is considered a region responsible for PKD, we recently reported that Prrt2 also regulates dopaminergic activity in the striatum, suggesting that Prrt2 functions not only in the cerebellum but also in the basal ganglia motor circuits. However, the relationship between neuronal cell types expressing Prrt2 and motor functions remains poorly understood. In this study, we determined the neurochemical types of Prrt2-positive neurons using immunofluorescence staining of mouse midbrain primary neurons and brain sections. Prrt2 was expressed mainly in glutamatergic and GABAergic neurons, but not in dopaminergic or cholinergic neurons. We found that Prrt2 was expressed preferentially in Vglut1-positive, rather than Vglut2-positive, cortical projection neurons and cerebellar granule cells, and in GABAergic medium spiny neurons of the basal ganglia, where Prrt2 was localized in axonal tracts and at or near presynaptic terminals. Taken together, we conclude that Prrt2 is variably expressed across excitatory and inhibitory neurons in motor-related neural circuits, where it might play more diverse roles in the regulation of neuronal excitability and voluntary movement.