Massively parallel in vivo Perturb-seq reveals cell-type-specific transcriptional networks in cortical development.

Leveraging AAVs' versatile tropism and labeling capacity, we expanded the scale of in vivo CRISPR screening with single-cell transcriptomic phenotyping across embryonic to adult brains and peripheral nervous systems. Through extensive tests of 86 vectors across AAV serotypes combined with a transposon system, we substantially amplified labeling efficacy and accelerated in vivo gene delivery from weeks to days. Our proof-of-principle in utero screen identified the pleiotropic effects of Foxg1, highlighting its tight regulation of distinct networks essential for cell fate specification of Layer 6 corticothalamic neurons. Notably, our platform can label >6% of cerebral cells, surpassing the current state-of-the-art efficacy at <0.1% by lentivirus, to achieve analysis of over 30,000 cells in one experiment and enable massively parallel in vivo Perturb-seq. Compatible with various phenotypic measurements (single-cell or spatial multi-omics), it presents a flexible approach to interrogate gene function across cell types in vivo, translating gene variants to their causal function.

Functional sensory circuits built from neurons of two species.

A central question for regenerative neuroscience is whether synthetic neural circuits, such as those built from two species, can function in an intact brain. Here, we apply blastocyst complementation to selectively build and test interspecies neural circuits. Despite approximately 10-20 million years of evolution, and prominent species differences in brain size, rat pluripotent stem cells injected into mouse blastocysts develop and persist throughout the mouse brain. Unexpectedly, the mouse niche reprograms the birth dates of rat neurons in the cortex and hippocampus, supporting rat-mouse synaptic activity. When mouse olfactory neurons are genetically silenced or killed, rat neurons restore information flow to odor processing circuits. Moreover, they rescue the primal behavior of food seeking, although less well than mouse neurons. By revealing that a mouse can sense the world using neurons from another species, we establish neural blastocyst complementation as a powerful tool to identify conserved mechanisms of brain development, plasticity, and repair.

Sexual coordination in a whole-brain map of prairie vole pair bonding.

Sexual bonds are central to the social lives of many species, including humans, and monogamous prairie voles have become the predominant model for investigating such attachments. We developed an automated whole-brain mapping pipeline to identify brain circuits underlying pair-bonding behavior. We identified bonding-related c-Fos induction in 68 brain regions clustered in seven major brain-wide neuronal circuits. These circuits include known regulators of bonding, such as the bed nucleus of the stria terminalis, paraventricular hypothalamus, ventral pallidum, and prefrontal cortex. They also include brain regions previously unknown to shape bonding, such as ventromedial hypothalamus, medial preoptic area, and the medial amygdala, but that play essential roles in bonding-relevant processes, such as sexual behavior, social reward, and territorial aggression. Contrary to some hypotheses, we found that circuits active during mating and bonding were largely sexually monomorphic. Moreover, c-Fos induction across regions was strikingly consistent between members of a pair, with activity best predicted by rates of ejaculation. A novel cluster of regions centered in the amygdala remained coordinated after bonds had formed, suggesting novel substrates for bond maintenance. Our tools and results provide an unprecedented resource for elucidating the networks that translate sexual experience into an enduring bond.

Sexual coordination in a whole-brain map of prairie vole pair bonding.

Sexual bonds are central to the social lives of many species, including humans, and monogamous prairie voles have become the predominant model for investigating such attachments. We developed an automated whole-brain mapping pipeline to identify brain circuits underlying pair-bonding behavior. We identified bonding-related c-Fos induction in 68 brain regions clustered in seven major brain-wide neuronal circuits. These circuits include known regulators of bonding, such as the bed nucleus of the stria terminalis, paraventricular hypothalamus, ventral pallidum, and prefrontal cortex. They also include brain regions previously unknown to shape bonding, such as ventromedial hypothalamus, medial preoptic area and the medial amygdala, but that play essential roles in bonding-relevant processes, such as sexual behavior, social reward and territorial aggression. Contrary to some hypotheses, we found that circuits active during mating and bonding were largely sexually monomorphic. Moreover, c-Fos induction across regions was strikingly consistent between members of a pair, with activity best predicted by rates of ejaculation. A novel cluster of regions centered in the amygdala remained coordinated after bonds had formed, suggesting novel substrates for bond maintenance. Our tools and results provide an unprecedented resource for elucidating the networks that translate sexual experience into an enduring bond.

A circuit from the locus coeruleus to the anterior cingulate cortex modulates offspring interactions in mice.

Social sensitivity to other individuals in distress is crucial for survival. The anterior cingulate cortex (ACC) is a structure involved in making behavioral choices and is influenced by observed pain or distress. Nevertheless, our understanding of the neural circuitry underlying this sensitivity is incomplete. Here, we reveal unexpected sex-dependent activation of ACC when parental mice respond to distressed pups by returning them to the nest ("pup retrieval"). We observe sex differences in the interactions between excitatory and inhibitory ACC neurons during parental care, and inactivation of ACC excitatory neurons increased pup neglect. Locus coeruleus (LC) releases noradrenaline in ACC during pup retrieval, and inactivation of the LC-ACC pathway disrupts parental care. We conclude that ACC maintains sex-dependent sensitivity to pup distress under LC modulation. We propose that ACC's involvement in parenting presents an opportunity to identify neural circuits that support sensitivity to the emotional distress of others.

High-throughput confocal airy beam oblique light-sheet tomography of brain-wide imaging at single-cell resolution.

Brain research is an area of research characterized by its cutting-edge nature, with brain mapping constituting a crucial aspect of this field. As sequencing tools have played a crucial role in gene sequencing, brain mapping largely depends on automated, high-throughput and high-resolution imaging techniques. Over the years, the demand for high-throughput imaging has scaled exponentially with the rapid development of microscopic brain mapping. In this paper, we introduce the novel concept of confocal Airy beam into oblique light-sheet tomography named CAB-OLST. We demonstrate that this technique enables the high throughput of brain-wide imaging of long-distance axon projection for the entire mouse brain at a resolution of 0.26 µm × 0.26 µm × 1.06 µm in 58 hours. This technique represents an innovative contribution to the field of brain research by setting a new standard for high-throughput imaging techniques.

Nephroprotective plant species used in traditional Mayan Medicine for renal-associated diseases.

ETHNOPHARMACOLOGICAL RELEVANCE: The prevalence of kidney disease has increased rapidly in recent years and has emerged as one of the leading causes of mortality worldwide. Natural products have been suggested as valuable nephroprotective agents due to their multi-target and synergistic effects on modulating important proteins involved in kidney injury. There is a large number of plant species that have been used traditionally for kidney-related conditions in Mesoamerican medicine by different cultural groups that could provide a valuable source of nephroprotective therapeutic candidates and could lead to potential drug discovery. AIM OF REVIEW: This review aims to provide an overview of the currently known efficacy of plant species used traditionally in Mesoamerica by Mayan groups to treat kidney-related conditions and to analyze the phytochemical, pharmacological, molecular, toxicological, and clinical evidence to contribute to public health efforts and for directing future research. METHODS: Primary sources of plant use reports for traditional kidney-related disorders in Mesoamerica were searched systematically from library catalogs, theses, and scientific databases (PubMed, Google Scholar; and Science Direct), and were filtered according to usage frequency in Mayan groups and plant endemism. The database of traditional plants was further analyzed based on associations with published reports of the phytochemical, pharmacological, molecular, toxicological, and clinical evidence. RESULTS: The most reported kidney-related conditions used traditionally in Mayan medicine involve reducing renal damage (a cultural interpretation that considers an inflammatory or infectious condition), cleaning or purifying the blood and kidney, reducing kidney pain, and eliminating kidney stones. A total of 208 plants used for kidney-related problems by 10 Mayan groups were found, representing 143 native species, where only 42 have reported pharmacological activity against kidney damage, mainly approached by in vitro and in vivo models of chemical- or drug-induced nephrotoxicity, diabetes nephropathy, and renal injury produced by hypertension. Nephroprotective effects are mainly mediated by reducing oxidative stress, inflammatory response, fibrosis mechanisms, and apoptosis in the kidney. The most common nephroprotective compounds associated with traditional Mayan medicine were flavonoids, terpenoids, and phenolic acids. The most widely studied traditional plants in terms of pharmacological evidence, bioactive compounds, and mechanisms of action, are Annona muricata L., Carica papaya L., Ipomoea batatas (L.) Lam., Lantana camara L., Sechium edule (Jacq.) Sw., Tagetes erecta L., and Zea mays L. Most of the plant species with reported pharmacological activity against kidney damage were considered safe in toxicological studies. CONCLUSION: Available pharmacological reports suggest that several herbs used in traditional Mayan medicine for renal-associated diseases may have nephroprotective effects and consistent pharmacological evidence, nephroprotective compounds, and mechanisms of action in different models of kidney injury. However, more research is required to fully understand the potential of traditional Mayan medicine in drug discovery given the limited ethnobotanical studies and data available for most species with regards to identification on bioactive components, pharmacological mechanisms, and the scarce number of clinical studies.

Quantitative relationship between cerebrovascular network and neuronal cell types in mice.

The cerebrovasculature and its mural cells must meet brain regional energy demands, but how their spatial relationship with different neuronal cell types varies across the brain remains largely unknown. Here we apply brain-wide mapping methods to comprehensively define the quantitative relationships between the cerebrovasculature, capillary pericytes, and glutamatergic and GABAergic neurons, including neuronal nitric oxide synthase-positive (nNOS+) neurons and their subtypes in adult mice. Our results show high densities of vasculature with high fluid conductance and capillary pericytes in primary motor sensory cortices compared with association cortices that show significant positive and negative correlations with energy-demanding parvalbumin+ and vasomotor nNOS+ neurons, respectively. Thalamo-striatal areas that are connected to primary motor sensory cortices also show high densities of vasculature and pericytes, suggesting dense energy support for motor sensory processing areas. Our cellular-resolution resource offers opportunities to examine spatial relationships between the cerebrovascular network and neuronal cell composition in largely understudied subcortical areas.

Medicinal plants used in traditional Mayan medicine for the treatment of central nervous system disorders: An overview.

ETHNOPHARMACOLOGICAL RELEVANCE: For thousands of years, different cultural groups have used and transformed natural resources for medicinal purposes focused on psychological or neurological conditions. Some of these are recognized as central nervous system (CNS) disorders and diseases, whereas other ethnopsychiatric interpretations are explained in culture-specific terms. In traditional Mayan medicine, several herbs have been part of treatments and rituals focused on cultural and ethnomedical concepts. AIM OF REVIEW: This study aims to provide a comprehensive overview of the medicinal plants used in Mesoamerica by traditional healers and Mayan groups to CNS disorders and associate the traditional use with demonstrated pharmacological evidence to establish a solid foundation for directing future research. METHODS: A systematic search for primary sources of plant use reports for traditional CNS-related remedies of Mesoamerica were obtained from library catalogs, thesis and scientific databases (PubMed, Scopus, Google Scholar; and Science Direct), and entered in a database with data analyzed in terms of the usage frequency, use by ethnic groups, plant endemism, and pharmacological investigation. RESULTS: A total of 155 plants used for ethnopsychiatric conditions in Mesoamerica by Mayan groups were found, encompassing 127 native species. Of these, only 49 native species have reported in vitro or in vivo pharmacological analyses. The most commonly reported ethnopsychiatric conditions are related to anxiety, depression, memory loss, epilepsy, and insomnia. The extent of the scientific evidence available to understand the pharmacological application for their use against CNS disorders varied between different plant species, with the most prominent evidence shown by Annona cherimola, Justicia pectoralis, J. spicigera, Mimosa pudica, Persea americana, Petiveria alliacea, Piper amalago, Psidium guajava, Tagetes erecta and T. lucida. CONCLUSION: Available pharmacological data suggest that different plant species used in traditional Mayan medicine may target the CNS, mainly related to GABA, serotonin, acetylcholine, or neuroprotective pathways. However, more research is required, given the limited data regarding mechanism of action at the preclinical in vivo level, identification of active compounds, scarce number of clinical studies, and the dearth of peer-reviewed studies.

Cellular anatomy of the mouse primary motor cortex.

An essential step toward understanding brain function is to establish a structural framework with cellular resolution on which multi-scale datasets spanning molecules, cells, circuits and systems can be integrated and interpreted1. Here, as part of the collaborative Brain Initiative Cell Census Network (BICCN), we derive a comprehensive cell type-based anatomical description of one exemplar brain structure, the mouse primary motor cortex, upper limb area (MOp-ul). Using genetic and viral labelling, barcoded anatomy resolved by sequencing, single-neuron reconstruction, whole-brain imaging and cloud-based neuroinformatics tools, we delineated the MOp-ul in 3D and refined its sublaminar organization. We defined around two dozen projection neuron types in the MOp-ul and derived an input-output wiring diagram, which will facilitate future analyses of motor control circuitry across molecular, cellular and system levels. This work provides a roadmap towards a comprehensive cellular-resolution description of mammalian brain architecture.

Oleoylethanolamide Delays the Dysfunction and Death of Purkinje Cells and Ameliorates Behavioral Defects in a Mouse Model of Cerebellar Neurodegeneration.

Oleoylethanolamide (OEA) is an endocannabinoid that has been proposed to prevent neuronal damage and neuroinflammation. In this study, we evaluated the effects of OEA on the disruption of both cerebellar structure and physiology and on the behavior of Purkinje cell degeneration (PCD) mutant mice. These mice exhibit cerebellar degeneration, displaying microtubule alterations that trigger the selective loss of Purkinje cells and consequent behavioral impairments. The effects of different doses (1, 5, and 10 mg/kg, i.p.) and administration schedules (chronic and acute) of OEA were assessed at the behavioral, histological, cellular, and molecular levels to determine the most effective OEA treatment regimen. Our in vivo results demonstrated that OEA treatment prior to the onset of the preneurodegenerative phase prevented morphological alterations in Purkinje neurons (the somata and dendritic arbors) and decreased Purkinje cell death. This effect followed an inverted U-shaped time-response curve, with acute administration on postnatal day 12 (10 mg/kg, i.p.) being the most effective treatment regimen tested. Indeed, PCD mice that received this specific OEA treatment regimen showed improvements in motor, cognitive and social functions, which were impaired in these mice. Moreover, these in vivo neuroprotective effects of OEA were mediated by the PPARα receptor, as pretreatment with the PPARα antagonist GW6471 (2.5 mg/kg, i.p.) abolished them. Finally, our in vitro results suggested that the molecular effect of OEA was related to microtubule stability and structure since OEA administration normalized some alterations in microtubule features in PCD-like cells. These findings provide strong evidence supporting the use of OEA as a pharmacological agent to limit severe cerebellar neurodegenerative processes.

Immunostimulatory Action of High-Content Active Arabinoxylan in Rice Bran.

Immunostimulatory activity comprises specific and nonspecific immune responses stimulated by internal and external factors. Arabinoxylan is well known for its immunostimulatory activity in vivo and in vitro, although the biological activities of arabinoxylan oligosaccharides depend on their structural features. In this study, we aimed to evaluate in vitro and in vivo the immunostimulatory activity of high-content active arabinoxylan (HCAA) obtained from rice bran through bioconversion by microorganisms and acid hydrolysis. Three microorganisms, Penicillium rocheforti, Aspergillus oryzae, and Pleurotus osteatus, and three different acid concentrations of hydrochloric acid (5, 10, and 20%) and acetic acid (25, 50, and 75%) were used for producing HCAA. HPLC analysis of arabinose and xylose content revealed that fermentation with P. rocheforti followed by hydrolysis with 5% hydrochloric acid was the most efficient to produce HCAA. GPC analysis of HCAA indicates that HCAA is a complex of various forms of saccharides and shows an average molecular weight of 625. Further, in vitro evaluation disclosed that exposure to HCAA (10-200 µg/mL) increased cell viability in mice splenic cells and RAW 264.7 cells. Additionally, exposure of mice to oral administration of HCAA (100 mg/kg) for 4-7 days increased lymphokine-activated killer (LAK)- and macrophage-mediated cytotoxic activity in cancer cells (YAC-1). Furthermore, in vitro exposure to HCAA and oral administrations in mice revealed increased interferon-γ (IFN-γ) and interleukin-10 (IL-10) protein expression through western blot analysis in RAW 264.7 cells and isolated splenic cells. Our results suggest that HCAA developed by bioconversion and acid hydrolysis may enhance immune responses in vivo and in vitro.

Biomarcadores traslacionales de modelos in vitro e in vivo de daño renal: Una perspectiva para abordar nefrotoxicidad desde múltiples factores etiológicos / Biomarcadores traslacionales de modelos in vitro e in vivo de daño renal: Una perspectiva para abordar nefrotoxicidad desde múltiples factores etiológicos

La prevalencia de enfermedad renal ha aumentado considerablemente en la última década y está previsto que crezca en los próximos años. Recientemente, diversos modelos se han utilizado para entender los procesos fisiopatológicos de daño renal y para la búsqueda de futuros candidatos farmacológicos. El objetivo de esta revisión es proporcionar una descripción de la evidencia actual de modelos in vitro e in vivo de nefrotoxicidad, nefropatía diabética y deshidratación, y los fundamentos de las principales vías de señalización fisiopatológicas, con el fin de proponer biomarcadores candidatos para futura investigación farmacológica. Actualmente, los roedores constituyen un pilar importante en estudios de daño renal, existiendo diferencias específicas según el estímulo nocivo, lo que sugiere considerar para un modelo relevante aspectos como especie, cepa, género y estructuras renales objetivo. Diversas estructuras renales se han complementado in vitro, principalmente a partir de líneas celulares, como del epitelio tubular, podocitos, células mesangiales glomerulares y conducto colector medular interno. Este enfoque se ha utilizado como complementario en modelos de nefrotoxicidad por exposición a aminoglucósidos (principalmente), deshidratación por cloruro de sodio hiperosmolar, y nefropatía diabética por medio de glucosa alta y productos derivados de glucólisis y glicación. Recientemente, estos modelos han mostrado similitud en diversas rutas de señalización celular, con algunos biomarcadores en común, entre múltiples causas de daño renal como el daño oxidativo, disfunción mitocondrial, procesos inflamatorios, desregulación de sistemas de defensa y sobrevivencia celular, y apoptosis. El enfoque en seleccionar biomarcadores relevantes contribuirá al diseño de estrategias terapéuticas de nefroprotectores sobre múltiples factores etiológicos.

The prevalence of kidney disease has increased considerably in the last decade and is expected to growth in the coming years. Recently, various models have been used to understand the pathophysiological processes of kidney damage and to search for future pharmacological candidates. The aim of this review is to provide a description of the current evidence of in vitro and in vivo models of nephrotoxicity, diabetic nephropathy and dehydration, and the foundations of the main pathophysiological signaling pathways, in order to propose candidate biomarkers for future drug discovery. Currently, rodents are an important pillar in studies of kidney damage, with specific differences depending on the noxious stimulus, which suggests considering aspects such as species, strain, gender and target structures for a relevant model. Several renal structures have been complemented through in vitro approaches, mainly using cell lines, such as the tubular epithelium, podocytes, glomerular mesangial cells and inner medullary collecting duct. These cells have been used as models of nephrotoxicity by exposure to aminoglycosides (mainly), dehydration by exposure to hyperosmolar sodium chloride, and diabetic nephropathy by exposure to high glucose and products derived from glycolysis and glycation. Recently, these models have shown common cell signaling pathways on multiple etiologies of kidney injury, sharing several biomarkers such as oxidative damage, mitochondrial dysfunction, inflammatory processes, dysregulation of defense systems and cell survival, and apoptosis. Approaching kidney injury based on the selection of relevant biomarkers will contribute to the design of therapeutic strategies for nephroprotection on multiple etiological factors.

Panax ginseng (Korea Red Ginseng) repairs diabetic sensorineural damage through promotion of the nerve growth factor pathway in diabetic zebrafish.

BACKGROUND: Diabetic sensorineural damage is a complication of the sensory neural system, resulting from long-term hyperglycemia. Red ginseng (RG) has shown efficacy for treatment of various diseases, including diabetes mellitus; however, there is little research about its benefit for treating sensorineural damage. Therefore, we aim to evaluate RG efficacy in alloxan-induced diabetic neuromast (AIDN) zebrafish. METHODS: In this study, we developed and validated an AIDN zebrafish model. To assess RG effectiveness, we observed morphological changes in live neuromast zebrafish. Also, zebrafish has been observed to have an ultrastructure of hair-cell cilia under scanning electron microscopy. Thus, we recorded these physiological traits to assess hair cell function. Finally, we confirmed that RG promoted neuromast recovery via nerve growth factor signaling pathway markers. RESULTS: First, we established an AIDN zebrafish model. Using this model, we showed via live neuromast imaging that RG fostered recovery of sensorineural damage. Damaged hair cell cilia were recovered in AIDN zebrafish. Furthermore, RG rescued damaged hair cell function through cell membrane ion balance. CONCLUSION: Our data suggest that RG potentially facilitates recovery in AIDN zebrafish, and its mechanism seems to be promotion of the nerve growth factor pathway through increased expression of topomyosin receptor kinase A, transient receptor potential channel vanilloid subfamily type 1, and mitogen-activated protein kinase phosphorylation.

Electrophysiological changes in auditory evoked potentials in rats with salicylate-induced tinnitus.

Early-response auditory evoked potentials (AEPs) in humans are significantly altered in tinnitus. These changes are closely related to that seen in animals, leading to new approaches to study tinnitus based on objective parameters. The purpose of this study was to characterize the AEPs in animals with tinnitus, by assessing early to late latency responses. For behavioral evaluation, rats were trained using positive reinforcement to press a lever in the presence of an auditory stimulus and to not press during silence. The auditory brainstem response (ABR), middle latency response (MLR) and auditory late latency response (LLR) were correlated to the false-positive responses (pressing the lever during silence), after oral administrations of Sodium Salicylate (SS, 350 mg/kg). In the present study, SS significantly increased the hearing thresholds and reduced ABR peak I amplitudes across the frequency range (4-32 kHz). In contrast, increased amplitudes were observed for several peaks in ABR, MLR, and LLR. Moreover, reduced ABR latencies in response to 8, 16 and 24 kHz tone bursts were observed after SS administration. Similarly, the central evaluation also revealed significantly reduced latencies in MLR and LLR during SS administration. In contrast, increased latencies were observed for ABR latencies in response to 32 kHz tone bursts, and at the P1-N1 component of LLR. Correlational analysis revealed that latencies and amplitudes of peaks II and IV (8 and 16 kHz) of ABR, and N2 latency and P2-N2 amplitude of LLR were associated with behavioral tinnitus. We suggest that AEPs can be used in the rat to evaluate the reduced sensory input and the increased central gain in SS-induced tinnitus, as well as reduced latencies (8-16 kHz) to distinguish between hearing loss and tinnitus.

Avocado Oil Extract Modulates Auditory Hair Cell Function through the Regulation of Amino Acid Biosynthesis Genes.

Sensorineural hearing loss (SNHL) is one of the most common causes of disability, affecting over 466 million people worldwide. However, prevention or therapy of SNHL has not been widely studied. Avocado oil has shown many health benefits but it has not yet been studied in regards to SNHL. Therefore, we aimed to investigate the efficacy of avocado oil on SNHL in vitro and in vivo and elucidate its mode of action. For the present study, we used enhanced functional avocado oil extract (DKB122). DKB122 led to recovery of otic hair cells in zebrafish after neomycin-induced otic cell damage. Also, DKB122 improved auditory sensory transmission function in a mouse model of noise induced-hearing loss and protected sensory hair cells in the cochlea. In addition, RNA sequencing was performed to elucidate the mechanism involved. KEGG pathway enrichment analysis of differentially expressed genes showed that DKB122 protected House Ear Institute-Organ of Corti 1 (HEI-OC1) cells against neomycin-related alterations in gene expression due to oxidative stress, cytokine production and protein synthesis.

Traditional oriental medicine for sensorineural hearing loss: Can ethnopharmacology contribute to potential drug discovery?

ETHNOPHARMACOLOGICAL RELEVANCE: In Traditional Oriental Medicine (TOM), the development of hearing pathologies is related to an inadequate nourishment of the ears by the kidney and other organs involved in regulation of bodily fluids and nutrients. Several herbal species have historically been prescribed for promoting the production of bodily fluids or as antiaging agents to treat deficiencies in hearing. AIM OF REVIEW: The prevalence of hearing loss has been increasing in the last decade and is projected to grow considerably in the coming years. Recently, several herbal-derived products prescribed in TOM have demonstrated a therapeutic potential for acquired sensorineural hearing loss and tinnitus. Therefore, the aims of this review are to provide a comprehensive overview of the current known efficacy of the herbs used in TOM for preventing different forms of acquired sensorineural hearing loss and tinnitus, and associate the traditional principle with the demonstrated pharmacological mechanisms to establish a solid foundation for directing future research. METHODS: The present review collected the literature related to herbs used in TOM or related compounds on hearing from Chinese, Korean, and Japanese herbal classics; library catalogs; and scientific databases (PubMed, Scopus, Google Scholar; and Science Direct). RESULTS: This review shows that approximately 25 herbal species and 40 active compounds prescribed in TOM for hearing loss and tinnitus have shown in vitro or in vivo beneficial effects for acquired sensorineural hearing loss produced by noise, aging, ototoxic drugs or diabetes. The inner ear is highly vulnerable to ischemia and oxidative damage, where several TOM agents have revealed a direct effect on the auditory system by normalizing the blood supply to the cochlea and increasing the antioxidant defense in sensory hair cells. These strategies have shown a positive impact on maintaining the inner ear potential, sustaining the production of endolymph, reducing the accumulation of toxic and inflammatory substances, preventing sensory cell death and preserving sensory transmission. There are still several herbal species with demonstrated therapeutic efficacy whose mechanisms have not been deeply studied and others that have been traditionally used in hearing loss but have not been tested experimentally. In clinical studies, Ginkgo biloba, Panax ginseng, and Astragalus propinquus have demonstrated to improve hearing thresholds in patients with sensorineural hearing loss and alleviated the symptoms of tinnitus. However, some of these clinical studies have been limited by small sample sizes, lack of an adequate control group or contradictory results. CONCLUSIONS: Current therapeutic strategies have proven that the goal of the traditional oriental medicine principle of increasing bodily fluids is a relevant approach for reducing the development of hearing loss by improving microcirculation in the blood-labyrinth barrier and increasing cochlear blood flow. The potential benefits of TOM agents expand to a multi-target approach on different auditory structures of the inner ear related to increased cochlear blood flow, antioxidant, anti-inflammatory, anti-apoptotic and neuroprotective activities. However, more research is required, given the evidence is very limited in terms of the mechanism of action at the preclinical in vivo level and the scarce number of clinical studies published.

Glycerides isolated from the aerial parts of Malva verticillata cause immunomodulation effects via splenocyte function and NK anti-tumor activity.

A preliminary study revealed that a 10 µg/mL n-BuOH fraction of Malva verticillata aerial parts significantly enhanced splenocyte proliferation and induced significant enhancement of natural-killer (NK) cell activity against tumor cells (YAC-1). This study was initiated to identify the principal components that exhibited these activities, and four glycerides were isolated through repeated SiO2 and ODS column chromatography. Structures of compounds 1-4 were determined to be (2S)-1-O-palmitoyl glyceride, (2S)-1-O-stearoyl glyceride, (2S)-1-O-linolenoyl glyceride, and (2S)-1,2-di-O-linoleoyl glyceride, respectively. Compounds 1-3 showed potential immune-enhancing activity in murine splenocyte and natural-killer (NK) cells at 10 µM. In contrast, compound 4 showed weak activity, indicating the monoacyl glycerides (1-3) are more effective than diacyl glyceride (4). Also, the longer the carbon number of the fatty acid in monoacyl glyceride, the better the activity, and the monoacyl glyceride including an unsaturated fatty acid (3) is more effective than the glycerides including the saturated fatty acids (1-2).

Cytoskeleton stability is essential for the integrity of the cerebellum and its motor- and affective-related behaviors.

The cerebellum plays a key role in motor tasks, but its involvement in cognition is still being considered. Although there is an association of different psychiatric and cognitive disorders with cerebellar impairments, the lack of time-course studies has hindered the understanding of the involvement of cerebellum in cognitive and non-motor functions. Such association was here studied using the Purkinje Cell Degeneration mutant mouse, a model of selective and progressive cerebellar degeneration that lacks the cytosolic carboxypeptidase 1 (CCP1). The effects of the absence of this enzyme on the cerebellum of mutant mice were analyzed both in vitro and in vivo. These analyses were carried out longitudinally (throughout both the pre-neurodegenerative and neurodegenerative stages) and different motor and non-motor tests were performed. We demonstrate that the lack of CCP1 affects microtubule dynamics and flexibility, defects that contribute to the morphological alterations of the Purkinje cells (PCs), and to progressive cerebellar breakdown. Moreover, this degeneration led not only to motor defects but also to gradual cognitive impairments, directly related to the progression of cellular damage. Our findings confirm the cerebellar implication in non-motor tasks, where the formation of the healthy, typical PCs structure is necessary for normal cognitive and affective behavior.

Trigonelline promotes auditory function through nerve growth factor signaling on diabetic animal models.

BACKGROUND: Protection of cochlear function and reconstruction of neuronal networks in damaged auditory sensory structures is crucial for therapeutic treatment of diabetic hearing loss. Nerve growth factor (NGF) has been used as a novel therapeutic target to protect against the neurodegenerative effects of Diabetes Mellitus (DM). PURPOSE: We aimed to evaluate the potential effect of trigonelline (TRG) on reducing auditory damage produced by DM using NGF as a potential marker. METHOD: Docking simulations were carried out using Autodock Vina software and visualized using Discovery Studio. Morphological analysis of hair cells and neuromasts was performed on alloxan-induced diabetic zebrafish by fluorescence and scanning electron microscopy. Blockage of NGF receptor phosphorylation with K-252a was used to evaluate TRG and NGF action. Further assessment of NGF by ELISA on a primary culture of spiral ganglion cells was performed as a marker of neuronal function on the hearing system. Finally, auditory function was assessed in LepR(db/db) mice using auditory brainstem response (ABR) and transient evoked otoacoustic emission (TEOAE) during 8 weeks. RESULTS: Docking simulations showed that TRG binds to the active site of NGF through molecular interactions with Lysine88 (Lys88) and Tyrosine52 (Tyr52). TRG treatment significantly reduced hair cell loss and neuromast damage in diabetic zebrafish (P < .05). Further evaluation revealed a significant increase in the number of neuromasts after NGF administration (P < .001). TRG and NGF action was suppressed during blockage of NGF receptor phosphorylation. Moreover, spiral ganglion cells revealed significant elevation on NGF values after TRG treatment (P < .05). In vivo evaluation of LepR(db/db) mice revealed a significant reduction in the auditory damage produced under diabetic progression, characterized by reduced ABR hearing threshold shifts and increased signal-to-noise ratio in TEOAE (P < .05). CONCLUSIONS: This study suggests that the enhanced hearing function produced by TRG may be mediated by NGF, providing a potential therapeutic strategy for diabetic hearing loss.