The plant natural product 2-methoxy-1,4-naphthoquinone stimulates therapeutic neural repair properties of olfactory ensheathing cells.

Olfactory ensheathing cells (OECs) are crucial for promoting the regeneration of the primary olfactory nervous system that occurs throughout life. Transplantation of OECs has emerged as a promising therapy for nervous system injuries, in particular for spinal cord injury repair. Functional outcomes in both animals and humans are, however, highly variable, primarily because it is difficult to rapidly obtain enough OECs for transplantation. Compounds which can stimulate OEC proliferation without changing the phenotype of the cells are therefore highly sought after. Additionally, compounds which can stimulate favourable cell behaviours such as migration and phagocytic activity are desirable. We conducted a medium-throughput screen testing the Davis open access natural product-based library (472 compounds) and subsequently identified the known plant natural product 2-methoxy-1,4-naphthoquinone as a stimulant of OEC viability. We showed that 2-methoxy-1,4-naphthoquinone: (i) strongly stimulates proliferation over several weeks in culture whilst maintaining the OEC phenotype; (ii) stimulates the phagocytic activity of OECs, and (iii) modulates the cell cycle. We also identified the transcription factor Nrf2 as the compound's potential molecular target. From these extensive investigations we conclude that 2-methoxy-1,4-naphthoquinone may enhance the therapeutic potential of OECs by stimulating proliferation prior to transplantation.

Evidence of axon connectivity across a spinal cord transection in rats treated with epidural stimulation and motor training combined with olfactory ensheathing cell transplantation.

Olfactory ensheathing cells (OECs) are unique glia that support axon outgrowth in the olfactory system, and when used as cellular therapy after spinal cord injury, improve recovery and axon regeneration. Here we assessed the effects of combining OEC transplantation with another promising therapy, epidural electrical stimulation during a rehabilitative motor task. Sprague-Dawley rats received a mid-thoracic transection and transplantation of OECs or fibroblasts (FBs) followed by lumbar stimulation while climbing an inclined grid. We injected pseudorabies virus (PRV) into hindlimb muscles 7 months post-injury to assess connectivity across the transection. Analyses showed that the number of serotonergic (5-HT) axons that crossed the rostral scar border and the area of neurofilament-positive axons in the injury site were both greater in OEC- than FB-treated rats. We detected PRV-labeled cells rostral to the transection and remarkable evidence of 5-HT and PRV axons crossing the injury site in 1 OEC- and 1 FB-treated rat. The axons that crossed suggested either axon regeneration (OEC) or small areas of probable tissue sparing (FB). Most PRV-labeled thoracic neurons were detected in laminae VII or X, and ~25% expressed Chx10, a marker for V2a interneurons. These findings suggest potential regeneration or sparing of circuits that connect thoracic interneurons to lumbar somatic motor neurons. Despite evidence of axonal connectivity, no behavioral changes were detected in this small-scale study. Together these data suggest that when supplemented with epidural stimulation and climbing, OEC transplantation can increase axonal growth across the injury site and may promote recovery of propriospinal circuitry.

The serrulatane diterpenoid natural products RAD288 and RAD289 stimulate properties of olfactory ensheathing cells useful for neural repair therapies.

Olfactory ensheathing cells (OECs) are being trialled for cell transplantation therapies for neural repair as they have unique properties which can enhance neuron regeneration. However, improvements in cell viability, proliferation and migration are needed to enhance therapeutic outcomes. Growth factors can enhance cell activity, but they can also induce side effects as they can act on numerous cell types. An alternative approach is to identify natural products (NPs) that more selectively activate specific cell functions. We have examined two pure NPs, 3-acetoxy-7,8-dihydroxyserrulat-14-en-19-oic acid (RAD288) and 3,7,8-trihydroxyserrulat-14-en-19-oic acid (RAD289) isolated from the Australian plant Eremophila microtheca. We determined that RAD288 and RAD289 stimulated the viability and proliferation of OECs in two-dimensional cultures and increased cell viability in three-dimensional spheroids. Both compounds also enhanced OEC-mediated phagocytosis of neural debris. However, only RAD288 stimulated migration of OECs, demonstrating that key structural changes to the compound can dramatically affect the resultant cellular action. In addition, cell-type specific action is highlighted by the result that neither compound stimulated the viability of Schwann cells which are a closely-related glial cell type. Therefore, these small molecules may have high potential for selective activation of specific therapeutically-useful activities of OECs for transplantation therapies to repair the nervous system.

Learning-Dependent and -Independent Enhancement of Mitral/Tufted Cell Glomerular Odor Responses Following Olfactory Fear Conditioning in Awake Mice.

Associative fear learning produces fear toward the conditioned stimulus (CS) and often generalization, the expansion of fear from the CS to similar, unlearned stimuli. However, how fear learning affects early sensory processing of learned and unlearned stimuli in relation to behavioral fear responses to these stimuli remains unclear. We subjected male and female mice expressing the fluorescent calcium indicator GCaMP3 in olfactory bulb mitral and tufted cells to a classical olfactory fear conditioning paradigm. We then used awake, in vivo calcium imaging to quantify learning-induced changes in glomerular odor responses, which constitute the first site of olfactory processing in the brain. The results demonstrate that odor-shock pairing nonspecifically enhances glomerular odor representations in a learning-dependent manner and increases representational similarity between the CS and nonconditioned odors, potentially priming the system toward generalization of learned fear. Additionally, CS-specific glomerular enhancements remain even when associative learning is blocked, suggesting two separate mechanisms lead to enhanced glomerular responses following odor-shock pairings.SIGNIFICANCE STATEMENT In the olfactory bulb (OB), odors are uniquely coded in a spatial map that represents odor identity, making the OB a unique model system for investigating how learned fear alters sensory processing. Classical fear conditioning causes fear of the conditioned stimulus (CS) and of neutral stimuli, known as generalization. Combining fear conditioning with fluorescent calcium imaging of OB glomeruli, we found enhanced glomerular responses of the CS as well as neutral stimuli in awake mice, which mirrors fear generalization. We report that CS and neutral stimuli enhancements are, respectively, learning-independent and learning-dependent. Together, these results reveal distinct mechanisms leading to enhanced OB processing of fear-inducing stimuli and provide important implications for altered sensory processing in fear generalization.

Odour-induced analgesia mediated by hypothalamic orexin neurons in mice.

Various folk remedies employ certain odorous compounds with analgesic effects. In fact, linalool, a monoterpene alcohol found in lavender extracts, has been found to attenuate pain responses via subcutaneous, intraperitoneal, intrathecal, and oral administration. However, the analgesic effects of odorous compounds mediated by olfaction have not been thoroughly examined. We performed behavioural pain tests under odourant vapour exposure in mice. Among six odourant molecules examined, linalool significantly increased the pain threshold and attenuated pain behaviours. Olfactory bulb or epithelium lesion removed these effects, indicating that olfactory sensory input triggered the effects. Furthermore, immunohistochemical analysis revealed that linalool activated hypothalamic orexin neurons, one of the key mediators for pain processing. Formalin tests in orexin neuron-ablated and orexin peptide-deficient mice showed orexinergic transmission was essential for linalool odour-induced analgesia. Together, these findings reveal central analgesic circuits triggered by olfactory input in the mammalian brain and support a potential therapeutic approach for treating pain with linalool odour stimulation.

Rauwolfia vomitoria inhibits olfaction and modifies olfactory bulb cells.

The rising cost of orthodox medication has endeared so many to the use of herbs for the management of neurological conditions. Rauwolfia vomitoria (RV) one of such herbs is a rainforest shrub whose parts are used locally in the management of psychiatry and other medical issues. Its usefulness though not in doubt is wrapped with adverse reports as its active constituents depletes brain monoamine and dopamine stores. This motivated this research on the effects of the root bark extract on olfaction and the olfactory bulb of adult Wistar rats. Eighteen adult Wistar rats (220g average) were divided into three groups (n=6); control (placebo), 200mg/kg and 400mg/kg RV root bark extract, respectively. The oral administration lasted for seven days and on day 8, test of olfaction was carried out and the animals immediately anaesthetized with ketamine hydrochloride (i.p.) and perfuse-fixed with 10% neutral buffered formalin. All the brains were processed for histology and immunoreactivity. Results showed loss of body weights and olfaction in the 200mg/kg and 400mg/kg RV groups. There was hypertrophy and atrophy of mitral cells respectively, in the 200mg/kg and 400mg/kg RV groups, while there was hyperplasia of cells in the internal granular and plexiform layers of both groups. There was decreased neuron specific enolase (NSE) and neurofilament (NF) expression in the 200mg/kg RV group, while NF and glial fibrillary acidic protein (GFAP) expression was decreased in the 400mg/kg RV group. However, NSE expression was enhanced in the 400mg/kg group, while GFAP expression was enhanced in the 200mg/kg RV group. These results suggest that these doses of RV affect olfaction and appetite, and stimulate adverse cellular changes in the olfactory bulb.

Cell migration in the developing rodent olfactory system.

The components of the nervous system are assembled in development by the process of cell migration. Although the principles of cell migration are conserved throughout the brain, different subsystems may predominantly utilize specific migratory mechanisms, or may display unusual features during migration. Examining these subsystems offers not only the potential for insights into the development of the system, but may also help in understanding disorders arising from aberrant cell migration. The olfactory system is an ancient sensory circuit that is essential for the survival and reproduction of a species. The organization of this circuit displays many evolutionarily conserved features in vertebrates, including molecular mechanisms and complex migratory pathways. In this review, we describe the elaborate migrations that populate each component of the olfactory system in rodents and compare them with those described in the well-studied neocortex. Understanding how the components of the olfactory system are assembled will not only shed light on the etiology of olfactory and sexual disorders, but will also offer insights into how conserved migratory mechanisms may have shaped the evolution of the brain.

Functional differentiation of cholinergic and noradrenergic modulation in a biophysical model of olfactory bulb granule cells.

Olfactory bulb granule cells are modulated by both acetylcholine (ACh) and norepinephrine (NE), but the effects of these neuromodulators have not been clearly distinguished. We used detailed biophysical simulations of granule cells, both alone and embedded in a microcircuit with mitral cells, to measure and distinguish the effects of ACh and NE on cellular and microcircuit function. Cholinergic and noradrenergic modulatory effects on granule cells were based on data obtained from slice experiments; specifically, ACh reduced the conductance densities of the potassium M current and the calcium-dependent potassium current, whereas NE nonmonotonically regulated the conductance density of an ohmic potassium current. We report that the effects of ACh and NE on granule cell physiology are distinct and functionally complementary to one another. ACh strongly regulates granule cell firing rates and afterpotentials, whereas NE bidirectionally regulates subthreshold membrane potentials. When combined, NE can regulate the ACh-induced expression of afterdepolarizing potentials and persistent firing. In a microcircuit simulation developed to investigate the effects of granule cell neuromodulation on mitral cell firing properties, ACh increased spike synchronization among mitral cells, whereas NE modulated the signal-to-noise ratio. Coapplication of ACh and NE both functionally improved the signal-to-noise ratio and enhanced spike synchronization among mitral cells. In summary, our computational results support distinct and complementary roles for ACh and NE in modulating olfactory bulb circuitry and suggest that NE may play a role in the regulation of cholinergic function.

Development of the thalamo-dorsal ventricular ridge tract in the Chinese soft-shelled turtle, Pelodiscus sinensis.

With the exception of that from the olfactory system, the vertebrate sensory information is relayed by the dorsal thalamus (dTh) to be carried to the telencephalon via the thalamo-telencephalic tract. Although the trajectory of the tract from the dTh to the basal telencephalon seems to be highly conserved among amniotes, the axonal terminals vary in each group. In mammals, thalamic axons project onto the neocortex, whereas they project onto the dorsal pallium and the dorsal ventricular ridge (DVR) in reptiles and birds. To ascertain the evolutionary development of the thalamo-telencephalic connection in amniotes, we focused on reptiles. Using the Chinese soft-shelled turtle (Pelodiscus sinensis), we studied the developmental course of the thalamic axons projecting onto the DVR. We found, during the developmental period when the thalamo-DVR connection forms, that transcripts of axon guidance molecules, including EphA4 and Slit2, were expressed in the diencephalon, similar to the mouse embryo. These results suggest that the basic mechanisms responsible for the formation of the thalamo-telencephalic tract are shared across amniote lineages. Conversely, there was a characteristic difference in the expression patterns of Slit2, Netrin1, and EphrinA5 in the telencephalon between synapsid (mammalian) and diapsid (reptilian and avian) lineages. This indicates that changes in the expression domains of axon guidance molecules may modify the thalamic axon projection and lead to the diversity of neuronal circuits in amniotes.

Histological study on effect of Nigella sativa on aged olfactory system of female albino rat.

Nigella sativa (NS) has wide-ranging healing properties, neuroprotective and antioxidant effects. Aging process is commonly associated with a decline in the chemical senses including smell. To detect a possible improvement effect of NS on the aging of the olfactory system we used 15 female albino rats that equally divided into three groups: group I (control adult), group II (control aged), group III (treated aged) received 40 mg/kg/day NS orally for two months. Specimens from the olfactory epithelium (OE), main olfactory bulb (MOB) and piriform cortex (PC) were processed for light and electron microscopy. Aging in OE revealed reduction in thickness, vacuolations, an increase in PAS reaction and lipofuscin autofluorescence. Aged MOB and PC exhibited a reduction in basophilia and accumulation of neurofibrillary tangles (NFTs) in mitral and pyramidal cells respectively. NS treatment improved the structure and the thickness of the OE and reduced the lipofuscin autofluorescence. It also attenuated the reduction in cytoplasmic basophilia and the accumulation of lipofuscin pigment and the NFTs in both mitral and pyramidal cells and the lipofuscin autofluorescence. These observations indicate that use of NS, could be of value in improving the structural changes of the peripheral and central main olfactory organs, which occurred in association with aging.

Sex hormone binding globulin in the rat olfactory system.

Ovarian steroids are known to act on the olfactory system. Their mode of action, however, is mostly unclear to date since nuclear receptors are lacking in sensory neurons. Here we used immunocytochemistry and RT-PCR to study expression and distribution of sex hormone binding globulin (SHBG) in the rat olfactory system. Single sensory cells in the olfactory mucosa and their projections in the olfactory bulb showed specific SHBG immunostaining as determined by double immunofluorescence with olfactory marker protein OMP. Larger groups of SHBG stained sensory cells occurred in the vomeronasal organ (VNO). A portion of the olfactory glomeruli in the accessory olfactory bulb showed large networks of SHBG positive nerve fibres. Some of the mitral cells showed SHBG immune fluorescence. RT-PCR revealed SHBG encoding mRNA in the olfactory mucosa, in the VNO and in the olfactory bulbs indicating intrinsic expression of the binding globulin. The VNO and its related projections within the limbic system are known to be sensitive to gonadal steroid hormones. We conclude that SHBG may be of functional importance for rapid effects of olfactory steroids on limbic functions including the control of reproductive behaviours through pheromones.

Hyperthermia-conditioned OECs serum-free-conditioned medium induce NSC differentiation into neuron more efficiently by the upregulation of HIF-1 alpha and binding activity.

BACKGROUND: Recent studies provide solid evidence for the importance to delineate the combined transplantation of neural stem cells (NSCs) and olfactory ensheathing cells (OECs) for the repair of central nervous system injury. One of the limitations of this approach is that the proportion of neurons differentiated from NSCs still remains at low level. Thus, how to induce NSCs to differentiate into neurons more efficiently by OECs is an attractive problem, which needs attention to be resolved. In the present study, we investigated the effects of hyperthermia-conditioned OEC-conditioned medium (OCM) on induction of NSCs into nerve cells. METHODS: The conditioned medium, named 37OCM, 40OCM, and 40OCM+R (treated with HIF-1a inhibitor), were collected after OECs had been incubated at 37°C or 40°C for 6 hr, followed by incubation at 37°C for 42 hr; then, these conditioned medium were collected and used for NSC induction. RESULTS: Results of the present study demonstrated for the first time that 6 hr of hyperthermia (40°C) could induce OECs upregulation and the expression of hypoxia-inducible factor-1 alpha (HIF-1α). Moreover, compared with 37OCM and 40OCM+R, 40OCM could induce NSCs differentiation into neurons more efficiently, and this phenomenon is associated with the upregulation of HIF-1α after hyperthermia conditioning. CONCLUSION: The data implied that the secretory protein in OECs-cultured medium and upregulation of HIF-1α expression and binding activity induced by hyperthermia-conditioned OECs have synergistic effect to induce NSCs differentiation into a neural lineage.

Greater excitability and firing irregularity of tufted cells underlies distinct afferent-evoked activity of olfactory bulb mitral and tufted cells.

Mitral and tufted cells, the two classes of principal neurons in the mammalian main olfactory bulb, exhibit morphological differences but remain widely viewed as functionally equivalent. Results from several recent studies, however, suggest that these two cell classes may encode complementary olfactory information in their distinct patterns of afferent-evoked activity. To understand how these differences in activity arise, we have performed the first systematic comparison of synaptic and intrinsic properties between mitral and tufted cells. Consistent with previous studies, we found that tufted cells fire with higher probability and rates and shorter latencies than mitral cells in response to physiological afferent stimulation. This stronger response of tufted cells could be partially attributed to synaptic differences, as tufted cells received stronger afferent-evoked excitation than mitral cells. However, differences in intrinsic excitability also contributed to the differences between mitral and tufted cell activity. Compared to mitral cells, tufted cells exhibited twofold greater excitability and peak instantaneous firing rates. These differences in excitability probably arise from differential expression of voltage-gated potassium currents, as tufted cells exhibited faster action potential repolarization and afterhyperpolarizations than mitral cells. Surprisingly, mitral and tufted cells also showed firing mode differences. While both cell classes exhibited regular firing and irregular stuttering of action potential clusters, tufted cells demonstrated a greater propensity to stutter than mitral cells. Collectively, stronger afferent-evoked excitation, greater intrinsic excitability and more irregular firing in tufted cells can combine to drive distinct responses of mitral and tufted cells to afferent-evoked input.

Intracranial transplant of olfactory ensheathing cells can protect both upper and lower motor neurons in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal disease that involves the degeneration of cortical and spinal motor neurons. Mutant SOD1(G93A) rats constitute a good animal model for this pathological condition. We have previously demonstrated that transplantation of neonatal olfactory ensheathing cells (OECs) into the dorsal funiculus of the spinal cord of mutant SOD1(G93A) transgenic rats increases the survival of spinal motor neurons and remyelinates the impaired axons through the pyramidal tract. In the present study, we examine whether intracranial cell implantation could also exert a similar effect on cortical motor neurons and on the lower motor neurons in the spinal cord. We injected OECs from the bulb of 7-day-old GFP green rats into the corona radiata of adult SOD1 mutant rats stereotaxically to observe any changes of the upper motor neurons as well as the lower motor neurons. We found that more motor neurons at both the motor cortices and ventral horns of the spinal cord survived in grafted ALS rats than in control rats. Prolonged survival and behavioral tests including a screen test, hind limb extension, rotarod, and gait control showed that the treated animals were better than the control group. This manuscript is published as part of the International Association of Neurorestoratology (IANR) supplement issue of Cell Transplantation.

A two-layer biophysical model of cholinergic neuromodulation in olfactory bulb.

Cholinergic inputs from the basal forebrain regulate multiple olfactory bulb (OB) functions, including odor discrimination, perceptual learning, and short-term memory. Previous studies have shown that nicotinic cholinergic receptor activation sharpens mitral cell chemoreceptive fields, likely via intraglomerular circuitry. Muscarinic cholinergic activation is less well understood, though muscarinic receptors are implicated in olfactory learning and in the regulation of synchronized oscillatory dynamics in hippocampus and cortex. To understand the mechanisms underlying cholinergic neuromodulation in OB, we developed a biophysical model of the OB neuronal network including both glomerular layer and external plexiform layer (EPL) computations and incorporating both nicotinic and muscarinic neuromodulatory effects. Our simulations show how nicotinic activation within glomerular circuits sharpens mitral cell chemoreceptive fields, even in the absence of EPL circuitry, but does not facilitate intrinsic oscillations or spike synchronization. In contrast, muscarinic receptor activation increases mitral cell spike synchronization and field oscillatory power by potentiating granule cell excitability and lateral inhibitory interactions within the EPL, but it has little effect on mitral cell firing rates and hence does not sharpen olfactory representations under a rate metric. These results are consistent with the theory that EPL interactions regulate the timing, rather than the existence, of mitral cell action potentials and perform their computations with respect to a spike timing-based metric. This general model suggests that the roles of nicotinic and muscarinic receptors in olfactory bulb are both distinct and complementary to one another, together regulating the effects of ascending cholinergic inputs on olfactory bulb transformations.

Setting the time course of inhibitory synaptic currents by mixing multiple GABA(A) receptor α subunit isoforms.

The kinetics of IPSCs influence many neuronal processes, such as the frequencies of oscillations and the duration of shunting inhibition. The subunit composition of recombinant GABA(A) receptors (GABA(A)Rs) strongly affects the deactivation kinetics of GABA-evoked currents. However, for GABAergic synapses, the relationship between subunit composition and IPSC decay is less clear. Here we addressed this by combining whole-cell recordings of miniature IPSCs (mIPSCs) and quantitative immunolocalization of synaptic GABA(A)R subunits. In cerebellar stellate, thalamic relay, and main olfactory bulb (MOB) deep short-axon cells of Wistar rats, the only synaptic α subunit was α1, and zolpidem-sensitive mIPSCs had weighted decay time constants (τ(w)) of 4-6 ms. Nucleus reticularis thalami neurons expressed only α3 as the synaptic α subunit and exhibited slow (τ(w) = 28 ms), zolpidem-insensitive mIPSCs. By contrast, MOB external tufted cells contained two α subunit types (α1 and α3) at their synapses. Quantitative analysis of multiple immunolabeled images revealed small within-cell, but large between-cell, variability in synaptic α1/α3 ratios. This corresponded to large cell-to-cell variability in the decay (τ(w) = 3-30 ms) and zolpidem sensitivity of mIPSCs. Currents evoked by rapid application of GABA to patches excised from HEK cells expressing different mixtures of α1 and α3 subunits displayed highly variable deactivation times that correlated with the α1/α3 cDNA ratio. Our results demonstrate that diversity in the decay of IPSCs can be generated by varying the expression of different GABA(A)R subunits that alone confer different decay kinetics, allowing the time course of inhibition to be tuned to individual cellular requirements.

In vitro effect of altering potassium concentration in artificial endolymph on apoptosis and ultrastructure features of olfactory bulb neural precursor cells.

Transplantation of neural stem cells (NSCs) into the cochlea to replace irreversibly damaged sensory epithelia is a potentially valuable remedy for hearing loss. Several mammalian stem cell lines are being successfully transplanted into, or migrated to, the endolymph (EL) fluids environment of the cochlea. However, the survival rate of transplanted cells is relatively low. This study focused on the effect of altering the potassium (K(+)) concentration of artificial EL on cell survival and apoptosis of olfactory bulb neural precursor cells (OB NPCs) in vitro. OB NPCs were prepared and placed in media for 24h, supplemented either with artificial EL, or artificial EL-like solutions of different K(+) concentrations. Survival, apoptotic features and ultrastructural changes in the cells are noted. Artificial EL-like solutions, especially with K(+) concentrations of 50mM or more, resulted in a series of necrotic or apoptotic events. Lower K(+) concentrations (30mM) decreased apoptosis and necrosis, improving the survival rate of cultured NPCs. Thus, it is conceivable that the external K(+) concentration in EL is a key environmental factor to regulate the survival of exogenous stem cells.

Transformation of odor representations in target areas of the olfactory bulb.

The brain generates coherent perceptions of objects from elementary sensory inputs. To examine how higher-order representations of smells arise from the activation of discrete combinations of glomeruli, we analyzed transformations of activity patterns between the zebrafish olfactory bulb and two of its telencephalic targets, Vv and Dp. Vv is subpallial whereas Dp is the homolog of olfactory cortex. Both areas lack an obvious topographic organization but perform complementary computations. Responses to different odors and their mixtures indicate that Vv neurons pool convergent inputs, resulting in broadened tuning curves and overlapping odor representations. Neuronal circuits in Dp, in contrast, produce a mixture of excitatory and inhibitory synaptic inputs to each neuron that controls action potential firing in an odor-dependent manner. This mechanism can extract information about combinations of molecular features from ensembles of active and inactive mitral cells, suggesting that pattern processing in Dp establishes representations of odor objects.

[Influence of acupuncture and moxibustion on proliferation and differentiation of neural stem cells in olfactory bulb of senescence-accelerated mouse P8].

This study was carried out to defect the effects of acupuncture and moxibustion therapy on the proliferation and differentiation of neural stem cells in the olfactory bulb of senescence-accelerated mouse P8. Immunofluorescence double staining methods(BrdU/Nestin, BrdU/MAP-2 and BrdU/beta-tubulin III) were used. The results show that both acupuncture and moxibustion could improve proliferation of neural stem cells, but only moxibustion could induce differentiation of neural stem cells into neuron.

Long-term functional restoration by neural progenitor cell transplantation in rat model of cognitive dysfunction: co-transplantation with olfactory ensheathing cells for neurotrophic factor support.

Neural progenitor cell transplantation has emerged as a promising approach for cell replacement therapy in the brain of neurodegenerative diseases. These are multipotent stem cells with self-renewal capabilities and can give rise to cells of all the three lineages of nervous system and can be maintained and differentiated to desirable neuronal subtypes in vitro with known trophic factors. However, like fetal cells, neural progenitor cells after differentiating to specific neuronal type also require continuous neurotrophic factor support for their long-term survival following transplantation. Recent reports suggest that olfactory ensheathing cells are capable of providing continuous neurotrophic factor to the transplanted neural progenitor cells for their long-term survival. In the present investigation, an attempt has been made to validate functional restoration in kainic acid lesioned rat model of cognitive dysfunction following co-transplantation of neural progenitor cells with olfactory ensheathing cells. Animals lesioned with kainic acid in CA3 subfield of hippocampal region were transplanted with neural progenitor cells, olfactory ensheathing cells or neural progenitor cells+olfactory ensheathing cells together. Twelve weeks post-transplantation functional restoration was assessed using neurobehavioral, neurochemical, and immunohistochemical approaches. Significant recovery in learning and memory (89%) was observed in co-transplanted group when compared to lesioned group. This was accompanied by significantly higher expression of choline acetyltransferase and restoration in cholinergic receptor binding in co-transplanted group (61%) over the animals transplanted either olfactory ensheathing cells or neural progenitor cells alone. Role of olfactory ensheathing cells in supplementing neurotrophic factors was further substantiated in vitro by pronounced differentiation of neural progenitor cells to choline acetyltransferase/acetylcholine esterase immunoreactive cells when co-cultured with olfactory ensheathing cells as compared to neural progenitor cells alone. The results strengthened the hypothesis that co-transplantation of olfactory ensheathing cells and neural progenitor cells may be a better approach for functional restoration in kainic acid induced rat model of cognitive dysfunction.