Surgical anatomy of innervation of the gallbladder in humans and Suncus murinus with special reference to morphological understanding of gallstone formation after gastrectomy

AIM： To clarify the innervation of human gallbladder,with special reference to morphological understanding of gallstone formation after gastrectomy.METHODS： The liver, gallbladder and surrounding structures were immersed in a 10 mg/L solution of alizarin red S in ethanol to stain the peripheral nerves in cadavers （n = 10）. Innervation in the areas was completely dissected under a binocular microscope. Similarly,innervation in the same areas of 10 Suncus murinus （S. murinus） was examined employing whole mount immunohistochemistry.RESULTS： Innervation of the gallbladder occurred predominantly through two routes. One was from the anterior hepatic plexus, the innervation occurred along the cystic arteries and duct. Invariably this route passed through the hepatoduodenal ligament. The other route was from the posterior hepatic plexus, the innervation occurred along the cystic duct ventrally. This route also passed through the hepatoduodenal ligament dorsally.Similar results were obtained in S. murinus.CONCLUSION： The route from the anterior hepatic plexus via the cystic artery and/or duct is crucial for preserving gallbladder innervation. Lymph node dissection specifically in the hepatoduodenal ligament may affect the incidence of gallstones after gastrectomy.Furthermore, the route from the posterior hepatic plexus via the common bile duct and the cystic duct to the gallbladder should not be disregarded. Preservation of the plexus may attenuate the incidence of gallstone formation after gastrectomy.

Adenomyosis uterine innervation in mice correlates to nerve growth factor expression,inflammation,and vascularization

BACKGROUND： Studies have shown that abnormal innervation is an important factor impacting occurrence and development of pathological pain in endometriosis. OBJECTIVE： To observe uterine innervation of adenomyosis mice and to analyze the cause of innervation changes due to nerve growth factor （NGF） expression, inflammation, and vascularization. DESIGN, TIME AND SETTING： This randomized, controlled, animal experiment was performed at the Research Institute of Obstetrics and Gynecology Hospital, and Central Laboratory of Zhongshan Hospital, Fudan University from March to December 2008. MATERIALS： Tamoxifen was provided by Fudan Forward, China. Rabbit anti-mouse NGF was purchased from Santa Cruz Corporation, USA; rabbit anti-protein gene product 9.5 （PGP9.5） and rabbit anti-substance P （SP） were purchased from Chemicon, USA. METHODS： A total of 40 newborn ICR mice were randomly assigned to adenomyosis model and control groups, with 20 animals in each group. Mice in the adenomyosis model group were orally administrated 2.7 μmol/kg tamoxifen on days 2-5 after birth, while the controls were not treated. MAIN OUTCOME MEASURES： Both uteri from all mice were harvested at days 135-145 after birth Expressions of polyclonal PGP9.5 and SP were immunohistochemically detected to demonstrate pan- and sensory nerve fibers. Microvessel density was quantified in the endometrium and myometrium using immunochemical staining for polyclonal rabbit anti-CD31, which stained vessels. Gene expression for NGF, high-affinity tyrosine kinase receptor （trkA）, p75 neuretrophin receptor （p75NTR）, bradykinin receptor-1 （BKR-1）, and 2 （BKR-2）, as well as substance P receptor （neurokininl receptor, NK1-R）, were detected by reverse transcription-polymerase chain reaction. NGF-13 protein expression was detected by Western blot analysis. RESULTS： More nerve fibers were stained with PGP9.5 in the endometrium and myometrium, and with SP in the endometrium, in adenomyosis mice compared with controls （P 〈 0.01 and P 〈 0.05）. Microvessel density in the myometrium of adenomyosis mice was significantly greater than the controls （P 〈 0.01）. In the uterus of adenomyosis mice, mRNA expression of NGF and its two receptors （trkA and p75 NTR）, BKR-1, and NK1-R, as well as protein expression of NGF-β, were greater than the control mice （P 〈 0.01 or P 〈 0.05）. CONCLUSION： Uterine innervation in the adenomyosis mice was increased compared with the controls. Moreover, NGF expression, inflammation, and vascularization, which have been shown to be impact factors of innervation, were abnormal in the uteri of adenomyosis mice.

Osteoblasts are inherently programmed to repel sensory innervation

Tissue innervation is a complex process controlled by the expression profile of signaling molecules secreted by tissue-resident cells that dictate the growth and guidance of axons.Sensory innervation is part of the neuronal network of the bone tissue with a defined spatiotemporal occurrence during bone development.Yet,the current understanding of the mechanisms regulating the map of sensory innervation in the bone tissue is still limited.Here,we demonstrated that differentiation of human mesenchymal stem cells to osteoblasts leads to a marked impairment of their ability to promote axonal growth,evidenced under sensory neurons and osteoblastic-lineage cells crosstalk.The mechanisms by which osteoblast lineage cells provide this nonpermissive environment for axons include paracrine-induced repulsion and loss of neurotrophic factors expression.We identified a drastic reduction of NGF and BDNF production and stimulation of Sema3A,Wnt4;and Shh expression culminating at late stage of OB differentiation.We noted a correlation between Shh expression profile,OB differentiation stages,and OB-mediated axonal repulsion.Blockade of Shh activity and signaling reversed the repulsive action of osteoblasts on sensory axons.Finally,to strengthen our model,we localized the expression of Shh by osteoblasts in bone tissue.Overall,our findings provide evidence that the signaling profile associated with osteoblast phenotype differentiating program can regulate the patterning of sensory innervation,and highlight osteoblast-derived Shh as an essential player in this cue-induced regulation.

Changes in corneal innervation and pain responses in fungal keratitis

AIM: To characterize changes in the cornea nerve and pain responses in fungal keratitis(FK).METHODS: A retrospective analysis of in vivo confocal microscopy images of 11 FK corneas was performed, and the results were compared with those for 11 normal corneas. Subbasal corneal nerves were analyzed for total nerve number, main nerve trunk number, branching patterns and tortuosity. C57 BL/6 mice were infected with Aspergillus fumigatus. Disease severity was determined through clinical scoring and slit lamp photography. Corneas were harvested at 1, 3, 5, and 7 d post infection(p.i.) and assessed for β III tubulin. Corneal mechanical sensitivity thresholds were detected by von Frey test. β-endorphin(β-EP) and μ receptor protein expression was detected through Western blotting.RESULTS: Total nerve number, main nerve trunk number, and nerve branching were significantly lower in FK patients than in controls, but tortuosity was not significantly different. In infected mice, subbasal nerve density decreased from 1 d p.i., reaching a minimum at 5 d p.i. Clinical scores rose at 1 d p.i., peaked at 3 d p.i., and decreased at 5 d p.i. Mechanical sensitivity thresholds showed the same trends. β-EP and μ receptor protein expression increased after infection.CONCLUSION: Corneal nerve density is lower in FK patients and Aspergillus fumigatus-infected mice than in controls. Pain sensitivity decreases with postinfection corneal ulcer aggravation. β-EP and μ receptor proteins are both upregulated in infected mouse corneas.

Role of intrahepatic innervation in regulating the activity of liver cells

Liver innervation comprises sympathetic,parasympathetic and peptidergic nerve fibers,organized as either afferent or efferent nerves with different origins and roles.Their anatomy and physiology have been studied in the past 30 years,with different results published over time.Hepatocytes are the main cell population of the liver,making up almost 80%of the total liver volume.The interaction between hepatocytes and nerve fibers is accomplished through a wealth of neurotransmitters and signaling pathways.In this short review,we have taken the task of condensing the most important data related to how the nervous system interacts with the liver and especially with the hepatocyte population,how it influences their metabolism and functions,and how different receptors and transmitters are involved in this complex process.

Does crossover innervation really affect the clinical outcome？ A comparison of outcome between unilateral and bilateral digital nerve repair

Digital nerve injuries are the mostly detected nerve injury in the upper extremity. However, since the clinical phenomenon of crossover innervation at some degree from uninjured digital nerve to the in- jured side occurs after digital nerve injuries is sustained, one could argue that this concept might even result in the overestimation of the outcome of the digital nerve repair. With this knowledge in mind, this study aimed to present novel, pure, focused and valuable clinical data by comparing the outcomes of bilateral and unilateral digital nerve repair. A retrospective review of 28 fingers with unilateral or bilateral digital nerve repair using end-to-end technique in 19 patients within 2 years was performed. Weber＇s two-point discrimination, sharp/dull discrimination, warm/cold sensation and Visual Analog Scale scoring were measured at final 12-month follow ups in all patients. There was no significant difference in recovery of sensibility after unilateral and bilateral digital nerve repairs. Though there is crossover innervation microscopically, it is not important in the clinical evaluation period. According to clinical findings from this study, crossover innervations appear to be negligible in the estimation of outcomes of digital neurorrhaphy.

Surgical anatomy of the innervation of pylorus in human and Suncus murinus, in relation to surgical technique for pylorus-preserving pancreaticoduodenectomy

AIM： To clarify the innervation of the antro-pyloric region in humans from a clinico-anatomical perspective. METHODS： The stomach, duodenum and surrounding structures were dissected in 10 cadavers, and immersed in a 10mg/L solution of alizarin red S in ethanol to stain the peripheral nerves. The distribution details were studied to confirm innervations in the above areas using a binocular microscope. Similarly, innervations in 10 Suncus murinus were examined using the method of whole-mount imm unohistochemistry. RESULTS： The innervation of the pyloric region in humans involved three routes： One arose from the anterior hepatic plexus via the route of the suprapyloric/supraduodenal branch of the right gastric artery; the second arose from the anterior and posterior gastric divisions, and the third originated from the posterior-lower region of the pyloric region, which passed via the infrapyloric artery or retroduodenal branches and was related to the gastroduodenal artery and right gastroepiploic artery. For Suncus murinus, results similar to those in humans were observed. CONCLUSION： There are three routes of innervation of the pyloric region in humans, wherein the route of the right gastric artery is most important for preserving pyloric region innervation. Function will be preserved by more than 80% by preserving the artery in pylorus-preserving pancreaticoduodenectomy （PPPD）. However, the route of the infrapyloric artery should not be disregarded. This route is related to several arteries （the right gastroepiploic and gastroduodenal arteries）, and the preserving of these arteries is advantageous for preserving pyloric innervation in PPPD. Concurrently, the nerves of Latarjet also play an important role in maintaining innervation of the antro-pyloric region in PPPD. This is why pyloric function is not damaged in some patients when the right gastric artery is dissected or damaged in PPPD.

A paracrine role for white thermogenic adipocytes in innervation: an evidence-based hypothesis

White adipose tissue(WAT) stores energy and also plays an important endocrine role in producing adipokines for communication with the peripheral and central nervous system. WAT consists of the major lipogenic unilocular adipocytes and the minor populations of beige and brite multilocular adipocytes. These multilocular adipocytes express thermogenic genes and have phenotypic similarity with thermogenic brown adipose tissue. According to a current paradigm, multilocular adipocytes have a thermogenic function in WAT. In this mini review, we discuss data revealing heterogeneity among multilocular cell subsets in WAT and their functions beyond thermogenesis. We propose a hypothetical neuroendocrine role for multilocular adipocytes subsets in the formation of adaptive sensory-sympathetic circuits between the central nervous system and adipose tissue, which activate lipolysis and thermogenesis in WAT in high energy demand situations.

Anatomical Evidence for the Neural Connection from the Emotional Brain to Autonomic Innervation in the Anterior Chamber Structures of the Eye

Objective Previous studies have shown that the autonomic nervous system(ANS),which can be affected by emotions,is important in the occurrence or progression of glaucoma.The autonomic innervation distributed in the anterior chamber(AC)structures might play an efferent role in the neural regulation of intraocular pressure(IOP).This study aimed to investigate the anatomic neural connection from the emotional brain to autonomic innervation in the AC.Methods A retrograde trans-multisynaptic pseudorabies virus encoded with an enhanced green fluorescent protein(PRV531)and non-trans-synaptic tracer FAST Dil were injected into the right eye of mice,respectively.Fluorescent localization in the emotional brain and preganglionic nuclei was studied.Five and a half days after PRV531 injection into the right AC,fluorescent signals were observed in several emotional brain regions,including the amygdala,agranular insular cortex,lateral septal nuclei,periaqueductal gray,and hypothalamus.Autonomic preganglionic nuclei,including Edinger-Westphal nucleus,superior salivatory nucleus,and intermediolateral nucleus,were labeled using PRV531.Results The sensory trigeminal nuclei were not labeled using PRV531.The fluorescence signals in the nuclei mentioned above showed bilateral distribution,primarily on the ipsilateral side.Seven days after injecting FAST Dil into the AC,we observed no FAST Dil-labeled neurons in the central nervous system.Conclusion Our results indicate a neural connection from the emotional brain to autonomic innervation in the AC,which provides anatomical support for the emotional influence of IOP via the ANS.

Regenerating nerve fiber innervation of extraocular muscles and motor functional changes following oculomotor nerve injuries at different sites

In the present study, the oculomotor nerves were sectioned at the proximal （subtentorial） and distal （superior orbital fissure） ends and repaired. After 24 weeks, vestibulo-ocular reflex evaluation confirmed that the regenerating nerve fibers following oculomotor nerve injury in the superior orbital fissure had a high level of specificity for innervating extraocular muscles. The level of functional recovery of extraocular muscles in rats in the superior orbital fissure injury group was remarkably superior over that in rats undergoing oculomotor nerve injuries at the proximal end （subtentorium）. Horseradish peroxidase retrograde tracing through the right superior rectus muscle showed that the distribution of neurons in the nucleus of the oculomotor nerve was directly associated with the injury site, and that crude fibers were badly damaged. The closer the site of injury of the oculomotor nerve was to the extraocular muscle, the better the recovery of neurological function was. The mechanism may be associated with the aberrant number of regenerated nerve fibers passing through the injury site.

Experimental Study of Corneal Innervation in Diabetic Mellitus

The morphology of corneal innervation was studied in rabbits with alloxan-induced diabetes.The experimental rabbits were killed after 6,8,10, 12weeks.Corneal innervation was studied by histochemical techniques and transimission electron microscopy.The results revealed that in diabetic rabbits,from 6 weeks,the density of nerve-nets was markedly sparser,the fibers were thinner and the activity of nerves was lower than those of the control group.Ultrastructural changes showed swelling of axons,irregular distribution of fibril and degeneration of mitochondria of varying degree.With the progress of disease the above changes were becoming more marked gradually.It may be assumed that these changes are related to hyperglycemia.

Is There an Incremental Prognostic Value of Evaluating Left Ventricular Dyssynchrony by Gated SPECT in Patients with Systolic Heart Failure and Altered Myocardial Sympathetic Innervation as Evaluated by Cardiac I-123 mIBG Imaging?

Background: Altered myocardial sympathetic innervation activity (AMSI) is known to be present in systolic heart failure patients (SHF) and recently SPECT imaging using I-123 mIBG heart to mediastinum (H/M) ratio <1.6 has been shown to predict MACE in the ADMIRE-HF trial. Left ventricular mechanical dyssynchrony (LVMD) is known to be present in a substantial number of SHF patients and has been studied mainly to guide CRT therapy. Recently gated SPECT has shown promise to provide an accurate assessment of LVMD. It remains unclear how the combination of AMSI and LVMD collectively affect clinical outcomes and other cardiovascular parameters. Objectives: The objectives are to examine the clinical characteristics and incremental prognostic value for MACE of LVMD determined by SPECT in SHF patients with or without abnormal cardiac MIBG uptake (H/M ratio < 1.6). Methods: Out of 30 SHF patients who participated from our institution in the ADMIRE-HF trial studying MIBG based AMSI, we included 22 patients with abnormal MIBG H/M ratio of <1.6. We performed gated SPECT LVMD analysis on these patients using the Emory Cardiac Toolbox. The 2 SPECT variables for LVMD assessed were histogram bandwidth and phase standard deviation both of which assess the extent of dispersion of LV activation during contraction as a marker of LVMD. Patients were followed up for a mean period of 6 years. The primary end point was mortality from any cause and secondary end point was heart failure admission or myocardial infarction or ICD shock. Results: 2 Groups were defined: Group A: n = 17 with H/M MIBG ratio < 1.6 and +LVMD and Group B, n = 5 H/M MIBG ratio &minus;LVMD. Baseline characteristics, cardiac risk factors and medications were comparable between both groups. LVEF was lower and RBBB was less common in Group A. There was no statistical difference in achievement of primary or secondary end points in the two groups including death heart failure readmissions, ICD shocks or MI. Conclusions: In our pilot study, we did not find definitive value of adding SPECT based LVMD to abnormal cardiac MIBG imaging in SHF patients with regards to predicting outcomes. Although our sample size is too small to make any definitive conclusions, it is possible that LVMD works independently through different pathways in the progression of SHF and hence may not necessarily add incremental value to AMSI determination using MIBG.

Crosstalk Between Peripheral Innervation and Pancreatic Ductal Adenocarcinoma

Pancreatic ductal adenocarcinoma(PDAC)is a highly aggressive lethal malignancy,characterized by late diagnosis,aggressive growth,and therapy resistance,leading to a poor overall prognosis.Emerging evidence shows that the peripheral nerve is an important non-tumor component in the tumor microenvironment that regulates tumor growth and immune escape.The crosstalk between the neuronal system and PDAC has become a hot research topic that may provide novel mechanisms underlying tumor progression and further uncover promising therapeutic targets.In this review,we highlight the mechanisms of perineural invasion and the role of various types of tumor innervation in the progression of PDAC,summarize the potential signaling pathways modulating the neuronal-cancer interaction,and discuss the current and future therapeutic possibilities for this condition.

Neural innervation in adipose tissue, gut, pancreas, and liver

Efficient communication between the brain and peripheral organs is indispensable for regulating physiological function and maintaining energy homeostasis. The peripheral nervous system (PNS) in vertebrates, consisting of the autonomic and somatic nervous systems, bridges the peripheral organs and the central nervous system (CNS). Metabolic signals are processed by both vagal sensory nerves and somatosensory nerves. The CNS receives sensory inputs via ascending nerves, serves as the coordination and integration center, and subsequently controls internal organs and glands via descending nerves. The autonomic nervous system consists of sympathetic and parasympathetic branches that project peripheral nerves into various anatomical locations to regulate the energy balance. Sympathetic and parasympathetic nerves typically control the reflexive and involuntary functions in organs. In this review article, we outline the innervation of adipose tissue, gut, pancreas, and liver, to illustrate the neurobiological basis of central-peripheral interactions. We emphasize the importance of understanding the functional atlas of neural control of energy metabolism, and more importantly, provide potential avenues for further research in this area.

A volar skin excisional wound model for in situ evaluation of multiple-appendage regeneration and innervation

Background:Promoting rapid wound healing with functional recovery of all skin appendages is the main goal of regenerative medicine.So far current methodologies,including the commonly used back excisional wound model(BEWM)and paw skin scald wound model,are focused on assessing the regeneration of either hair follicles(HFs)or sweat glands(SwGs).How to achieve de novo appendage regeneration by synchronized evaluation of HFs,SwGs and sebaceous glands(SeGs)is still challenging.Here,we developed a volar skin excisional wound model(VEWM)that is suitable for examining cutaneous wound healing with multiple-appendage restoration,as well as innervation,providing a new research paradigm for the perfect regeneration of skin wounds.Methods:Macroscopic observation,iodine-starch test,morphological staining and qRT-PCR anal-ysis were used to detect the existence of HFs,SwGs,SeGs and distribution of nerve fibres in the volar skin.Wound healing process monitoring,HE/Masson staining,fractal analysis and behavioral response assessment were performed to verify that VEWM could mimic the pathological process and outcomes of human scar formation and sensory function impairment.Results:HFs are limited to the inter-footpads.SwGs are densely distributed in the footpads,scattered in the IFPs.The volar skin is richly innervated.The wound area of the VEWM at 1,3,7 and 10 days after the operation is respectively 89.17%±2.52%,71.72%±3.79%,55.09%±4.94%and 35.74%±4.05%,and the final scar area accounts for 47.80%±6.22%of the initial wound.While the wound area of BEWM at 1,3,7 and 10 days after the operation are respectively 61.94%±5.34%,51.26%±4.89%,12.63%±2.86%and 6.14%±2.84%,and the final scar area accounts for 4.33%±2.67%of the initial wound.Fractal analysis of the post-traumatic repair site for VEWM vs human was performed:lacunarity values,0.040±0.012 vs 0.038±0.014;fractal dimen-sion values,1.870±0.237 vs 1.903±0.163.Sensory nerve function of normal skin vs post-traumatic repair site was assessed:mechanical threshold,1.05±0.52 vs 4.90 g±0.80;response rate to pin-prick,100%vs 71.67%±19.92%,and temperature threshold,50.34◦C±3.11◦C vs 52.13◦C±3.54◦C.Conclusions:VEWM closely reflects the pathological features of human wound healing and can be applied for skin multiple-appendages regeneration and innervation evaluation.

3D anatomy of autonomic innervations in immune organs of a non-human primate and the human

Direct neural inputs to immune organs have been observed for decades,with their functions in neuroimmune regulation being increasingly appreciated.However,the current knowledge of such neural structures,particularly those in primate immune organs,remains incomplete.In this study,we comprehensively assessed the 3D anatomy of autonomic(i.e.,sympathetic and parasympathetic)innervations in the immune organs of the rhesus macaque monkey and the human for the first time.Aided with the advanced technique of whole-tissue immunolabeling and lightsheet fluorescence imaging,we revealed the densely organized sympathetic architecture in the parenchyma of the adult monkey and human spleens.On the other hand,only sparse,if any,sympathetic inputs were observed inside the lymph nodes,Peyer's patches,or thymus.In contrast,there were minimal parasympathetic innervations in the parenchyma of these examined immune organs.Together,this work has documented the unique patterns of autonomic innervations in different immune organs of a non-human primate and the human,serving as an essential reference for future research on neuroimmune regulation in the field.

Correlation between Cholinergic Innervation, Autophagy, and Etiopathology of Benign Prostatic Hyperplasia

Background： Whether cholinergic innervations and/or autophagy have a role in the etiopathology of benign prostatic hyperplasia （BPH） is still unknown. This study aimed to investigate the role of cholinergic innervation and autophagy in the etiopathology of BPH. Methods： Male, 13-week-old spontaneous hypertension rats （spontaneous BPH animal model） were divided into three groups： an experimental group （EG, n = 24）, a control group （CG, n = 24）, and a normal control group （NC, n = 10）. The EG animals were intragastrically injected with tolterodine （3.5 mg/kg, twice a day）, CG animals were intragastrically injected with physiological saline, and the NC animals did not receive any treatment. Rats were sacrificed every 4 weeks, and the prostatic gross morphological changes, wet weight/body weight （ww/bw）, dry weight/wet weight （dw/ww）, histological changes, ultrastructural changes, and LC3 immunohistochemistry were continuously observed and compared. Results： The gross morphological and ww/bw changes in the three groups were similar at every stage. The dw/ww （mg/mg） values of the EG at week 17, 21, 25, and 29 were 0.1478 ±0.0034, 0.1653 ± 0.0036, 0.1668 ± 0.0045, and 0.1755±0.0034, respectively, and the CG values were 0.1511 ±0.0029, 0.1734± 0.0020, 0.1837 ±0.0052, and 0.1968 ± 0.0045, respectively. The difference between EG and CG for dw/ww showed statistical significance after 21 weeks of age （week 21： P = 0.016, week 25： P = 0.008, and week 29： P = 0.001）. Both EG and CG, prostatic glandular epithelial cell proliferation, and secretory function improved with age, but in EG, these improvements were slower than those in CG, and all the differences were statistically significant after 21 weeks. An increasing number of autophagosomes in the prostatic glandular cell cytoplasm, attenuation of LC3-I immunohistochemical staining, enhancement of LC3-II staining, and the ratio of LC3-1I/LC3-1 staining were all progressive in both groups, but the rate of change in EG was faster than that in CG, and these differences gained statistical significance after 25 weeks. Comparisons with regard to the above indexes between CG and NC showed no statistical significance at any stage. Conclusions： Cholinergic innervations and activation of autophagy appear to have important functions in the etiopathology of BPH. Drug-mediated blockade of cholinergic innervations could delay the physiopathology processes. Moreover, overactivation of autophagy may also play an important role in this delay.

Differential Innervation of Secretory Coils and Ducts in Human Eccrine Sweat Glands

Background： Previous studies demonstrate that eccrine sweat glands are innervated by both cholinergic and adrenergic nerves. However, it is still unknown whether the secretory coils and ducts ofeccrine sweat glands are equally innervated by the sympathetic nerve fibers. To welt understand the mechanisms on sweat secretion and reabsorption, the differential innervation of secretory coils and ducts in human eccrine sweat glands was investigated in the study. Methods： From June 2016 to June 2017, six human skins were fixed, paraffin-embedded, and cut into 5 μm-thick sections, followed by costaining for nerve fiber markers protein gene product 9.5 （PGP 9.5）, tyrosine hydroxylase （TH） and vasoactive intestinal peptide （VIP）, and eccrine sweat gland markers K7, S100P, and K14 by combining standard immunofluorescence with tyramide signal amplification （IF-TSA）. Stained sections were observed under the microscope, photographed, and analyzed. Results： The fluorescent signals of PGP 9.5, TH, and VIP were easily visualized, by 1F-TSA, as circular patterns surrounding eccrine sweat glands, but only PGP 9.5 could be observed by standard IF. The IF-TSA method is more sensitivity than standard IF in detecting antigens expressed at low levels. PGP 9.5, TH, and VIP appeared primarily surrounding the secretory coils and sparsely surrounding the sweat ducts. Conclusion： Sweat secretion is mainly controlled by autonomic nerves whereas sweat reabsorption is less affected by nerve activity.

Innervation frequency response of sounding muscles in Mingming cicada

In cicadas the songs are produced by vibrations of the sounding membrane driven by sounding muscles. The innervations of sounding movements are synchro-controlling type for some species and asynchro-controlling type for other species. The contracting movements of sounding muscles on both sides are synchronous for some species and alternate for other species, and the contracting frequencies are from several decade to several hundred and generally below 500 Hz. On the basis of studies on the song property and innervation of sound production in the Mingnfing cicada, the innervation frequency-response of movements in sounding muscles is studied in this note. The results provide the important basis for expounding the sounding mechanism of vari-toned songs in cicadas.

Development of glutamatergic innervation during maturation of adult-born neurons

The dentate gyrus is the entrance of the hippocampal formation and a primary target of excitatory afferents from the entorhinal cortex that carry spatial and sensory information. Mounting evidence suggests that continual adult neurogenesis contributes to appropriate processing of cortical information. The ongoing integration of adult born neurons dynamically modulates connectivity of the network, potentially contributing to dentate cognitive function. Here we review the current understanding of how glutamatergie innervation develops during the progression of adult-born neuron maturation. Summarizing the developmental stages of dentate neurogenesis, we also demonstrate that new neurons at an immature stage of maturation begin to process afferent activity from both medial and lateral entorhinal cortices.