Injection Of Nuclei Research Articles

Magnetic Resonance Imaging-Guided Frameless Stereotactic Injections of the Bilateral Cerebellar Dentate Nuclei in Nonhuman Primates: Technical Note.

Nonhuman primates (NHPs) are important preclinical models for evaluating therapeutics because of their anatomophysiological similarities to humans, and can be especially useful for testing new delivery targets. With the growing promise of cell and gene therapies for the treatment of neurological diseases, it is important to ensure the accurate and safe delivery of these agents to target structures in the brain. However, a standard guideline or method has not been developed for stereotactic targeting in NHPs. In this article, we describe the safe use of a magnetic resonance imaging-guided frameless stereotactic system to target bilateral cerebellar dentate nuclei for accurate, real-time delivery of viral vector in NHPs. Seventeen rhesus macaques (Macaca mulatta) underwent stereotactic surgery under real-time MRI guidance using the ClearPoint® system. Bilateral cerebellar dentate nuclei were targeted through a single parietal entry point with a transtentorial approach. Fifty microliters of contrast-impregnated infusate was delivered to each dentate nucleus, and adjustments were made as necessary according to real-time MRI monitoring of delivery. Perioperative clinical outcomes and postoperative volumes of distribution were recorded. All macaques underwent bilateral surgery successfully. Superficial pin site infection occurred in 4/17 (23.5%) subjects, which resolved with antibiotics. Two episodes of transient neurological deficit (anisocoria and unilateral weakness) were recorded, which did not require additional postoperative treatment and resolved over time. Volume of distribution of infusate achieved satisfactory coverage of target dentate nuclei, and only 1 incidence (2.9%) of cerebrospinal fluid penetration was recorded. Mean volume of distribution was 161.22 ± 39.61 mm3 (left, 173.65 ± 48.29; right, 148.80 ± 23.98). MRI-guided frameless stereotactic injection of bilateral cerebellar dentate nuclei in NHPs is safe and feasible. The use of this technique enables real-time modification of the surgical plan to achieve adequate target coverage and can be readily translated to clinical use.

The Roles of Periaqueductal Gray and Dorsal Raphe Nucleus Dopaminergic Systems in the Mechanisms of Thermal Hypersensitivity and Depression in Mice

Depression and thermal hypersensitivity share pathogenic features and symptomology, but their pathophysiologic interactions have not been fully elucidated. Dopaminergic systems in the ventrolateral periaqueductal gray (vlPAG) and dorsal raphe nucleus have been implicated in these conditions due to their antinociception and antidepression effects, although their specific roles and underlying mechanisms remain obscure. In this study, chronic unpredictable mild stress (CMS) was used to induce depression-like behaviors and thermal hypersensitivity in C57BL/6J (wild-type) or dopamine transporter promoter mice to establish a mouse model of pain and depression comorbidity. Microinjections of quinpirole, a dopamine D2 receptor agonist, up-regulated D2 receptor expression in dorsal raphe nucleus and reduced depressive behaviors and thermal hypersensitivity with CMS, while dorsal raphe nucleus injections of JNJ-37822681, an antagonist of D2 receptors, had the reciprocal effect on dopamine D2 receptor expression and behaviors. Moreover, using a chemical genetics approach to activate or inhibit dopaminergic neurons in vlPAG ameliorated or exacerbated depression-like behaviors and thermal hypersensitivity, respectively, in dopamine transporter promoter-Cre CMS mice. Collectively these results demonstrated the specific role of vlPAG and dorsal raphe nucleus dopaminergic systems in the regulation of pain and depression comorbidity in mice. PerspectiveThe current study provides insights into the complex mechanisms underlying thermal hypersensitivity induced by depression, and the findings suggest that pharmacological and chemogenetic modulation of dopaminergic systems in the vlPAG and dorsal raphe nucleus may be a promising therapeutic strategy to simultaneously mitigate pain and depression.

Nucleation and cavitation inception in high Reynolds number shear layers

The influence of nucleation on cavitation inception in a high Reynolds number shear layer in the wake of a backward-facing step was experimentally investigated in a water tunnel. The flow was investigated for two nuclei populations: the one naturally occurring in the water and for the water artificially seeded with monodisperse nuclei. Incipient events were observed to form in stretched quasi-streamwise vortices. The collapse of an incipient cavity resulted in a microbubble cloud dispersed into the shear layer and the step re-circulation zone. These microbubbles, generally larger than those naturally occurring in the water, act as preferential sites for re-nucleation, triggering the formation of developed cavitation. This phenomenon rendered statistical characterization of cavitation inception impractical for the natural nuclei population. The re-nucleation issue was addressed by seeding the flow with a population of large monodisperse nuclei, with a critical pressure higher than that of cavitation products. Spatial distribution of the nuclei within the seeded plume was characterized using a volumetric measurement based on Mie-scattering imaging. The ability to discern individual incipient events enabled examination of the effect of cavitation number and the nuclei injection rate on the inception event rate. The event rate was found to follow a power law with cavitation number and vary linearly with the injection rate. Mapping of spatial distribution of cavitation susceptibility was obtained by combining the spatial distributions of incipient events and nuclei concentration. The current work provides a valuable dataset for the development of computational tools for modeling of cavitation inception in nucleated flows.

First Application of Mass Measurements with the Rare-RI Ring Reveals the Solar r-Process Abundance Trend at A=122 and A=123.

The Rare-RI Ring (R3) is a recently commissioned cyclotronlike storage ring mass spectrometer dedicated to mass measurements of exotic nuclei far from stability at Radioactive Isotope Beam Factory (RIBF) in RIKEN. The first application of mass measurement using the R3 mass spectrometer at RIBF is reported. Rare isotopes produced at RIBF-^{127}Sn, ^{126}In, ^{125}Cd, ^{124}Ag, ^{123}Pd-were injected in R3. Masses of ^{126}In, ^{125}Cd, and ^{123}Pd were measured whereby the mass uncertainty of ^{123}Pd was improved. This is the first reported measurement with a new storage ring mass spectrometry technique realized at a heavy-ion cyclotron and employing individual injection of the preidentified rare nuclei. The latter is essential for the future mass measurements of the rarest isotopes produced at RIBF. The impact of the new ^{123}Pd result on the solar r-process abundances in a neutron star merger event is investigated by performing reaction network calculations of 20 trajectories with varying electron fraction Y_{e}. It is found that the neutron capture cross section on ^{123}Pd increases by a factor of 2.2 and β-delayed neutron emission probability, P_{1 n}, of ^{123}Rh increases by 14%. The neutron capture cross section on ^{122}Pd decreases by a factor of 2.6 leading to pileup of material at A=122, thus reproducing the trend of the solar r-process abundances. The trend of the two-neutron separation energies (S_{2n}) was investigated for the Pd isotopic chain. The new mass measurement with improved uncertainty excludes large changes of the S_{2n} value at N=77. Such large increase of the S_{2n} values before N=82 was proposed as an alternative to the quenching of the N=82 shell gap to reproduce r-process abundances in the mass region of A=112-124.

Descending Dopaminergic Inputs to Reticulospinal Neurons Promote Locomotor Movements.

Meso-diencephalic dopaminergic neurons are known to modulate locomotor behaviors through their ascending projections to the basal ganglia, which in turn project to the mesencephalic locomotor region, known to control locomotion in vertebrates. In addition to their ascending projections, dopaminergic neurons were found to increase locomotor movements through direct descending projections to the mesencephalic locomotor region and spinal cord. Intriguingly, fibers expressing tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine synthesis, were also observed around reticulospinal neurons of lampreys. We now examined the origin and the role of this innervation. Using immunofluorescence and tracing experiments, we found that fibers positive for dopamine innervate reticulospinal neurons in the four reticular nuclei of lampreys. We identified the dopaminergic source using tracer injections in reticular nuclei, which retrogradely labeled dopaminergic neurons in a caudal diencephalic nucleus (posterior tuberculum [PT]). Using voltammetry in brain preparations isolated in vitro, we found that PT stimulation evoked dopamine release in all four reticular nuclei, but not in the spinal cord. In semi-intact preparations where the brain is accessible and the body moves, PT stimulation evoked swimming, and injection of a D1 receptor antagonist within the middle rhombencephalic reticular nucleus was sufficient to decrease reticulospinal activity and PT-evoked swimming. Our study reveals that dopaminergic neurons have access to command neurons that integrate sensory and descending inputs to activate spinal locomotor neurons. As such, our findings strengthen the idea that dopamine can modulate locomotor behavior both via ascending projections to the basal ganglia and through descending projections to brainstem motor circuits.SIGNIFICANCE STATEMENT Meso-diencephalic dopaminergic neurons play a key role in modulating locomotion by releasing dopamine in the basal ganglia, spinal networks, and the mesencephalic locomotor region, a brainstem region that controls locomotion in a graded fashion. Here, we report in lampreys that dopaminergic neurons release dopamine in the four reticular nuclei where reticulospinal neurons are located. Reticulospinal neurons integrate sensory and descending suprareticular inputs to control spinal interneurons and motoneurons. By directly modulating the activity of reticulospinal neurons, meso-diencephalic dopaminergic neurons control the very last instructions sent by the brain to spinal locomotor circuits. Our study reports on a new direct descending dopaminergic projection to reticulospinal neurons that modulates locomotor behavior.

Quipazine Elicits Swallowing in the Arterially Perfused Rat Preparation: A Role for Medullary Raphe Nuclei?

Pharmacological neuromodulation of swallowing may represent a promising therapeutic option to treat dysphagia. Previous studies suggested a serotonergic control of swallowing, but mechanisms remain poorly understood. Here, we investigated the effects of the serotonergic agonist quipazine on swallowing, using the arterially perfused working heart-brainstem (in situ) preparation in rats. Systemic injection of quipazine produced single swallows with motor patterns and swallow-breathing coordination similar to spontaneous swallows, and increased swallow rate with moderate changes in cardiorespiratory functions. Methysergide, a 5-HT2 receptor antagonist, blocked the excitatory effect of quipazine on swallowing, but had no effect on spontaneous swallow rate. Microinjections of quipazine in the nucleus of the solitary tract were without effect. In contrast, similar injections in caudal medullary raphe nuclei increased swallow rate without changes in cardiorespiratory parameters. Thus, quipazine may exert an excitatory effect on raphe neurons via stimulation of 5-HT2A receptors, leading to increased excitability of the swallowing network. In conclusion, we suggest that pharmacological stimulation of swallowing by quipazine in situ represents a valuable model for experimental studies. This work paves the way for future investigations on brainstem serotonergic modulation, and further identification of neural populations and mechanisms involved in swallowing and/or swallow-breathing interaction.

Functional Connectivity Between the Trigeminal Main Sensory Nucleus and the Trigeminal Motor Nucleus.

The present study shows new evidence of functional connectivity between the trigeminal main sensory (NVsnpr) and motor (NVmt) nuclei in rats and mice. NVsnpr neurons projecting to NVmt are most highly concentrated in its dorsal half. Their electrical stimulation induced multiphasic excitatory synaptic responses in trigeminal MNs and evoked calcium responses mainly in the jaw-closing region of NVmt. Induction of rhythmic bursting in NVsnpr neurons by local applications of BAPTA also elicited rhythmic firing or clustering of postsynaptic potentials in trigeminal motoneurons, further emphasizing the functional relationship between these two nuclei in terms of rhythm transmission. Biocytin injections in both nuclei and calcium-imaging in one of the two nuclei during electrical stimulation of the other revealed a specific pattern of connectivity between the two nuclei, which organization seemed to critically depend on the dorsoventral location of the stimulation site within NVsnpr with the most dorsal areas of NVsnpr projecting to the dorsolateral region of NVmt and intermediate areas projecting to ventromedial NVmt. This study confirms and develops earlier experiments by exploring the physiological nature and functional topography of the connectivity between NVsnpr and NVmt that was demonstrated in the past with neuroanatomical techniques.

Some new hints on cosmic-ray propagation from AMS-02 nuclei spectra

In this work, we considered 2 schemes (a high-rigidity break in primary source injections and a high-rigidity break in diffusion coefficient) to reproduce the newly released AMS-02 nuclei spectra (He, C, N, O, Li, Be, and B) when the rigidity larger than 50 GV. The fitting results show that current data set favors a high-rigidity break at ∼ 325 GV in diffusion coefficient rather than a break at ∼ 365 GV in primary source injections. Meanwhile, the fitted values of the factors to rescale the cosmic-ray (CR) flux of secondary species/components after propagation show us that the secondary flux are underestimated in current propagation model. It implies that we might locate in a slow diffusion zone, in which the CRs propagate with a small value of diffusion coefficient compared with the averaged value in the galaxy. Another hint from the fitting results show that extra secondary CR nuclei injection may be needed in current data set. All these new hints should be paid more attention in future research.

7T MRI Predicts Amelioration of Neurodegeneration in the Brain after AAV Gene Therapy

GM1 gangliosidosis (GM1) is a fatal neurodegenerative lysosomal storage disease that occurs most commonly in young children, with no effective treatment available. Long-term follow-up of GM1 cats treated by bilateral thalamic and deep cerebellar nuclei (DCN) injection of adeno-associated virus (AAV)-mediated gene therapy has increased lifespan to 8 years of age, compared with an untreated lifespan of ~8 months. Due to risks associated with cerebellar injection in humans, the lateral ventricle was tested as a replacement route to deliver an AAVrh8 vector expressing feline β-galactosidase (β-gal), the defective enzyme in GM1. Treatment via the thalamus and lateral ventricle corrected storage, myelination, astrogliosis, and neuronal morphology in areas where β-gal was effectively delivered. Oligodendrocyte number increased, but only in areas where myelination was corrected. Reduced AAV and β-gal distribution were noted in the cerebellum with subsequent increases in storage, demyelination, astrogliosis, and neuronal degeneration. These postmortem findings were correlated with endpoint MRI and magnetic resonance spectroscopy (MRS). Compared with the moderate dose with which most cats were treated, a higher AAV dose produced superior survival, currently 6.5 years. Thus, MRI and MRS can predict therapeutic efficacy of AAV gene therapy and non-invasively monitor cellular events within the GM1 brain.

Effects of Intracerebroventricular and Intra-Arcuate Nucleus Injection of Ghrelin on Pain Behavioral Responses and Met-Enkephalin and β-Endorphin Concentrations in the Periaqueductal Gray Area in Rats.

Ghrelin is an endogenous ligand for orphan growth hormone secretagogue receptors. Ghrelin receptors have been found in central nervous system (CNS) areas responsible for pain modulation and transmission. This study investigated the effects of intracerebroventricular (ICV) and intra-arcuate nucleus (ARC) injection of ghrelin on pain behavioral responses and levels of β-endorphin (β-EP) and met-enkephalin (MENK) in the periaqueductal gray area (PAG) during the formalin test in rats. Thirty-five male rats were studied in five groups. Ghrelin was injected into the left lateral ventricle (ICV, 5 µL) or into the ARC (1 µL). After 15 min, formalin (2.5%) was subcutaneously injected into the left hind paw. Behavioral nociceptive scores were recorded for 60 min. MENK and β-EP were collected by microdialysis in the PAG and determined by high-performance liquid chromatography (HPLC). ICV and ARC injection of ghrelin significantly reduced pain in all phases of the formalin test (p < 0.001). Dialysate concentrations of MENK and β-EP in the PAG increased in all the phases (p < 0.01). In conclusion, the present study shows that the ARC nucleus and the endogenous opioid system are involved in ghrelin-induced pain modulation.

Ultrahigh energy cosmic rays and neutrinos from light nuclei composition

The baryonic mass composition of ultrahigh energy ($\gtrsim 10^{18}$ eV) cosmic rays (UHECRs) at injection accompanied by their interactions on universal photon backgrounds during propagation directly governs the UHECR flux on the Earth. Secondary neutrinos and photons produced in these interactions serve as crucial astrophysical messengers of UHECR sources. A modeling of the latest data obtained by the Pierre Auger Observatory (PAO) suggests a mixed element composition of UHECRs with the sub-ankle spectrum being explained by a different class of sources than the super-ankle region ($> 10^{18.7}$ eV). In this work, we obtain two kinds of fit to the UHECR spectrum -- one with a single population of sources comprising of $^1$H and $^2$He, over an energy range commencing at $\approx 10^{18}$ eV -- another for a mixed composition of representative nuclei $^1$H, $^4$He, $^{14}$N and $^{28}$Si at injection, for which a fit is obtained from above $\approx 10^{18.7}$ eV. In both cases, we consider the source emissivity evolution to be a simple power-law in redshift. We test the credibility of H+He composition by varying the source properties over a wide range of values and compare the results to that obtained for H+He+N+Si composition, using the Monte Carlo simulation tool CRPropa 3. The secondary electrons and photons are propagated using the cosmic ray transport code DINT. We place limits on the source spectral index, source evolution index and cutoff rigidity of the source population in each case by fitting the UHECR spectrum. Cosmogenic neutrino fluxes can further constrain the abundance fraction and maximum source redshift in case of light nuclei injection model.

ARCiS framework for exoplanet atmospheres

Context. Understanding of clouds is instrumental in interpreting current and future spectroscopic observations of exoplanets. Modeling clouds consistently is complex, since it involves many facets of chemistry, nucleation theory, condensation physics, coagulation, and particle transport. Aims. We aim to develop a simple physical model for cloud formation and transport, efficient and versatile enough that it can be used, in modular fashion for parameter optimization searches of exoplanet atmosphere spectra. In this work we present the cloud model and investigate the dependence of key parameters as the cloud diffusivity K and the nuclei injection rate Σn. on the planet’s observational characteristics. Methods. The transport equations are formulated in 1D, accounting for sedimentation and diffusion. The grain size is obtained through a moment method. For simplicity, only one cloud species is considered and the nucleation rate is parametrized. From the resulting physical profiles we simulate transmission spectra covering the visual to mid-IR wavelength range. Results. We apply our models toward KCl clouds in the atmosphere of GJ1214 b and toward MgSiO3 clouds of a canonical hot-Jupiter. We find that larger K increases the thickness of the cloud, pushing the τ = 1 surface to a lower pressure layer higher in the atmosphere. A larger nucleation rate also increases the cloud thickness while it suppresses the grain size. Coagulation is most important at high Σn. and low K. We find that the investigated combinations of K and Σn. greatly affect the transmission spectra in terms of the slope at near-IR wavelength (a proxy for grain size), the molecular features seen at approximately μm (which disappear for thick clouds, high in the atmosphere), and the 10 μm silicate feature, which becomes prominent for small grains high in the atmosphere. Conclusions. Clouds have a major impact on the atmospheric characteristics of hot-Jupiters, and models as those presented here are necessary to reveal the underlying properties of exoplanet atmospheres. The result of our hybrid approach – aimed to provide a good balance between physical consistency and computational efficiency – is ideal toward interpreting (future) spectroscopic observations of exoplanets.

DEVELOPMENT OF ANTIHAIL ROCKET WITH REACTION LIFTOFF

Hail-suppression activities are undertaken in many countries to prevent the devastating effects of hailstorms. Among various hail-suppression techniques there is overseeding, where the hail formation process is influenced by seeding the clouds with artificial nuclei. An injection of artificial nuclei into supercooled parts of the clouds can be performed by small low-altitude rockets which carry a pyrotechnic substance impregnated with meteorological reagent. The reagent produces nuclei during burning of pyrotechnic substance and injects these nuclei directly into the cloud region where the hail develops and thus prevents hail formation. This paper reports on the development of a new generation of antihail rockets, where a rocket (which consists of a motor and a container with a pyrotechnic substance) is launched from its transport tube, i.e., the transport tube represents the launcher at the same time. In addition to motor thrust, the rocket escapes the launch tube at the expense of exhaust gases produced by the motor (hot-launch system). As the rocket launch velocity increases in this way, its operation range is extended and the surface wind sensitivity of the rocket trajectory is reduced. Thereby the total mass of the propellant and the diameter of the rocket are reduced, for the same maximum altitude and mass of the reagent, as compared to the rockets propelled during launching by thrust force only. This also ensures better performance, easy handling, and reliability of use.

Cell Injection Millirobot Development and Evaluation in Microfluidic Chip.

We propose an innovative design of millirobot, which can achieve donor cell suction, delivery, and injection in a mammalian oocyte on a microfluidic chip. The millirobot body contains a hollow space that produces suction and ejection forces for the injection of cell nuclei using a nozzle at the tip of the robot. Specifically, a controller changes the hollow volume by balancing the magnetic and elastic forces of a membrane along with the motion of stages in the XY plane. A glass capillary attached to the tip of the robot contains a nozzle that is able to absorb and inject cell nuclei. The millirobot provides three degrees of freedom and generates micronewton forces. We demonstrate the effectiveness of the proposed millirobot through an experiment of the absorption and ejection of 20-µm particles from the nozzle using magnetic control in a microfluidic chip.

Establishment of a Valuable Mimic of Alzheimer's Disease in Rat Animal Model by Intracerebroventricular Injection of Composited Amyloid Beta Protein.

Alzheimer's disease (AD) is an irreversible, progressive brain disease that slowly destroys memory and is accompanied by neuron loss and structure change. With the increase of AD patients worldwide, the pathology and treatment of the disease has become a focus in the International Pharmaceutical Industry. Thus, the establishment of the animal model to mimic AD in the laboratory is of great importance.Here, we describe a detailed protocol for establishing a mimic of AD in a rat animal model though intracerebroventricular injection of amyloid beta protein 25-35 (Aβ25-35) combined with aluminum trichloride (AlCl3) and anterodorsal thalamic nucleus injection of recombinant human transforming growth factor-β1 (RHTGF-β1) to rats. The related markers of AD were measured, including: spatial memory, neuronal structure and substructure, neuronal Aβ, and neurofibrillary tangles (NFT) production. This rat model demonstrates spatial memory impairment, neuronal structure and substructure pathological changes, neuron intracellular Aβ burden, and NFT aggregation, and provides a close mimic of the neuronal structure and function disorder to that of clinical AD patients. Thus, the presented AD rat modelprovides a valuable in vivo tool for exploring neuronal function, neuronal pathology, and drug screening of AD.

Establishment of a Valuable Mimic of Alzheimer's Disease in Rat Animal Model by Intracerebroventricular Injection of Composited Amyloid Beta Protein

Alzheimer's disease (AD) is an irreversible, progressive brain disease that slowly destroys memory and is accompanied by neuron loss and structure change. With the increase of AD patients worldwide, the pathology and treatment of the disease has become a focus in the International Pharmaceutical Industry. Thus, the establishment of the animal model to mimic AD in the laboratory is of great importance. Here, we describe a detailed protocol for establishing a mimic of AD in a rat animal model though intracerebroventricular injection of amyloid beta protein 25-35 (Aβ25-35) combined with aluminum trichloride (AlCl3) and anterodorsal thalamic nucleus injection of recombinant human transforming growth factor-β1 (RHTGF-β1) to rats. The related markers of AD were measured, including: spatial memory, neuronal structure and substructure, neuronal Aβ, and neurofibrillary tangles (NFT) production. This rat model demonstrates spatial memory impairment, neuronal structure and substructure pathological changes, neuron intracellular Aβ burden, and NFT aggregation, and provides a close mimic of the neuronal structure and function disorder to that of clinical AD patients. Thus, the presented AD rat modelprovides a valuable in vivo tool for exploring neuronal function, neuronal pathology, and drug screening of AD.

Cosmic ray and neutrino emission from gamma-ray bursts with a nuclear cascade

Aim. We discuss neutrino and cosmic ray emission from gamma-ray bursts (GRBs) with the injection of nuclei, where we take into account that a nuclear cascade from photodisintegration can fully develop in the source. Our main objective is to test whether recent results from the IceCube and the Pierre Auger Observatory can be accommodated within the paradigm that GRBs are the sources of ultra-high-energy cosmic rays (UHECRs). Methods. We simulate this scenario in a combined source-propagation model. While our key results are obtained using an internal shock model of the source, we discuss how the secondary emission from a GRB shell can be interpreted in terms of other astrophysical models. Results. We demonstrate that the expected neutrino flux from GRBs weakly depends on the injection composition for the same injection spectra and luminosities, which implies that prompt neutrinos from GRBs can efficiently test the GRB-UHECR paradigm even if the UHECRs are nuclei. We show that the UHECR spectrum and composition, as measured by the Pierre Auger Observatory, can be self-consistently reproduced. In an attempt to describe the energy range including the ankle, we find tension with the IceCube bounds from the GRB stacking analyses. In an alternative scenario, where only the UHECRs beyond the ankle originate from GRBs, the requirement for a joint description of cosmic ray and neutrino observations favors lower luminosities, which does not correspond to the typical expectation from γ-ray observations.

Ketamine-Induced Prefrontal Serotonin Release Is Mediated by Cholinergic Neurons in the Pedunculopontine Tegmental Nucleus.

BackgroundKetamine rapidly elicits antidepressive effects in humans and mice in which serotonergic activity is involved. Although α4β2 nicotinic acetylcholine receptor (α4β2 nAChR) in the dorsal raphe nucleus plays a key role in the ketamine-induced prefrontal serotonin release, the source of cholinergic afferents, and its role is unclear.MethodsPrefrontal serotonin levels after ketamine injection were measured by microdialysis in rats. Electrolytic lesion of pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus was made with constant direct current.ResultsBilateral lesion of the pedunculopontine tegmental nucleus, but not laterodorsal tegmental nucleus, attenuated prefrontal serotonin release induced by systemic ketamine. Intra-pedunculopontine tegmental nucleus, but not intra-laterodorsal tegmental nucleus ketamine perfusion, increased prefrontal serotonin release. This increase was attenuated by intra-dorsal raphe nucleus injection of dihydro-β-erythroidine, an α4β2 nAChR antagonist, or NBQX, an AMPA receptor antagonist.ConclusionsThese results suggest the ketamine-induced serotonin release in medial prefrontal cortex is mediated by cholinergic neurons projecting from pedunculopontine tegmental nucleus to dorsal raphe nucleus via α4β2 nAChRs.

Effect and Mechanism of ARC Nucleus Injection of CART on Insulin Secretion in Islet β Cells of Type 2 Diabetic Rats

Effect and Mechanism of ARC Nucleus Injection of CART on Insulin Secretion in Islet β Cells of Type 2 Diabetic Rats

Interspecies somatic cell nuclear transfer in Asiatic cheetah using nuclei derived from post-mortem frozen tissue in absence of cryo-protectant and in vitro matured domestic cat oocytes

Recent accomplishments in the field of somatic cell nuclear transfer (SCNT) hold tremendous promise to prevent rapid loss of animal genetic resources using ex situ conservation technology. Most of SCNT studies use viable cells for nuclear transfer into recipient oocytes. However, preparation of live cells in extreme circumstances, in which post-mortem material of endangered/rare animals is improperly retained frozen, is difficult, if not impossible. This study investigated the possibility of interspecies-SCNT (iSCNT) in Asiatic cheetah (Acinonyx jubatus venaticus), a critically endangered subspecies, using nuclei derived from frozen tissue in absence of cryo-protectant at −20 °C and in vitro matured domestic cat oocytes. No cells growth was detected in primary culture of skin and tendon pieces or following culture of singled cells prepared by enzymatic digestion. Furthermore, no live cells were detected following differential viable staining and almost all cells had ruptured membrane. Therefore, direct injection of donor nuclei into enucleated cat oocytes matured in vitro was carried out for SCNT experiments. Early signs of nuclear remodeling were observed as early as 2 h post-iSCNT and significantly increased at 4 h post-iSCNT. The percentages of iSCNT reconstructs that cleaved and developed to 4–16 cell and morula stages were 32.3 ± 7.3, 18.2 ± 9.8 and 5.9 ± 4.3%, respectively. However, none of the iSCNT reconstructs developed to the blastocyst stage. When domestic cat somatic and oocytes were used for control SCNT and parthenogenetic activation, the respective percentages of oocytes that cleaved (51.3 ± 13.9 and 77.3 ± 4.0%) and further developed to the blastocyst stage (11.3 ± 3.3 and 16.8 ± 3.8%) were comparable. In summary, this study demonstrated that enucleated cat oocytes can partially remodel and reactivate non-viable nuclei of Asiatic cheetah and support its reprogramming back to the embryonic stage. To our knowledge, this is the first report of iSCNT in cheetah using non-viable frozen cells.