J.W. Jahng

Mesolimbic dopaminergic activity responding to acute stress is blunted in adolescent rats that experienced neonatal maternal separation

Neonatal maternal separation (MS), stressful experience early in life, leads to the development of depression-like behaviors in the offspring later in life. This study was conducted to define the neural basis of depression-like behaviors observed in our MS model. Sprague-Dawley pups were separated from dam for 3 h daily during the first 2 weeks of birth (MS) or left undisturbed (NH). All pups were sacrificed on postnatal day 41 with/without 1 h of restraint stress. Restraint stress significantly increased c-Fos expression in the nucleus accumbens (NAcb) of NH pups, but not in MS. In NH pups, restraint stress increased dopamine levels not only in the NAcb but also in the midbrain dopamine neurons; however, these increases were not observed in MS. Gene expression of tyrosine hydroxylase (TH) in the ventral tegmental area (VTA) was increased by acute restraint in NH pups, but not in MS pups. The raphe serotonin level was lower in MS than in NH, and not significantly changed by acute restraint neither in NH nor in MS. Results reveal that experience of neonatal MS may lead to a long-term suppression in the mesolimbic dopamine system of the offspring later in life, in which an epigenetic control may be implicated, such as suppressed gene expression of TH in the midbrain. We conclude that a decreased activity of the mesolimbic dopamine system may play a role in the pathophysiology of depression-like behaviors by neonatal MS, in addition to a decreased serotonin level in the raphe nucleus.

The arcuate NPY, POMC, and CART expressions responding to food deprivation are exaggerated in young female rats that experienced neonatal maternal separation.

This study was conducted to examine the effect of neonatal maternal separation on the hypothalamic feeding peptides expression in young female offspring. Sprague-Dawley pups were separated from dam for 3h daily during PND 1-14 (MS), or left undisturbed except routine cage cleaning (NH). Weanling female pups were housed in group and the arcuate mRNA levels of neuropeptide Y (NPY), proopiomelanocortin (POMC), and cocaine-amphetamine regulated transcript (CART) were examined at two months of age with or without food deprivation. The basal arcuate expression levels of these peptides did not differ between NH and MS group. However, a 48 h of food deprivation significantly increased NPY mRNA level, and decreased POMC and CART, in the arcuate nucleus of MS females, but not in NH females. Fasting-induced elevation of the plasma corticosterone tended to be greater in MS group than in NH, but the basal levels did not differ between the groups. Plasma leptin levels were decreased in MS females compared with NH, and food deprivation significantly suppressed the leptin levels both in NH and MS groups. Results suggest that MS experience may increase stress vulnerability in female rats and exaggerate the feeding peptides expression in the arcuate nucleus responding to metabolic stress food deprivation.

Repeated oral administration of capsaicin increases anxiety-like behaviours with prolonged stress-response in rats

This study was conducted to examine the psycho-emotional effects of repeated oral exposure to capsaicin, the principal active component of chili peppers. Each rat received 1 mL of 0.02% capsaicin into its oral cavity daily, and was subjected to behavioural tests following 10 daily administrations of capsaicin. Stereotypy counts and rostral grooming were significantly increased, and caudal grooming decreased, in capsaicin-treated rats during the ambulatory activity test. In elevated plus maze test, not only the time spent in open arms but also the percent arm entry into open arms was reduced in capsaicin-treated rats compared with control rats. In forced swim test, although swimming duration was decreased, struggling increased in the capsaicin group, immobility duration did not differ between the groups. Repeated oral capsaicin did not affect the basal levels of plasma corticosterone; however, the stress-induced elevation of plasma corticosterone was prolonged in capsaicin treated rats. Oral capsaicin exposure significantly increased c-Fos expression not only in the nucleus tractus of solitarius but also in the paraventricular nucleus. Results suggest that repeated oral exposure to capsaicin increases anxiety-like behaviours in rats, and dysfunction of the hypothalamic-pituitary-adrenal axis may play a role in its pathophysiology.

Co-treatment effect of pulsed electromagnetic field (PEMF) with human dental pulp stromal cells and FK506 on the regeneration of crush injured rat sciatic nerve

Purpose. The purpose of this study was to determine whether crush injured rat sciatic nerve could be benefit from pulsed electromagnetic field (PEMF) combined with human dental pulp stromal cells (hDPSCs), with FK506 (Tacrolimus) for immune suppression and neuropromotion. Materials and Methods. Male Sprague-Dawley rats (200–250 g, 6 week old) were distributed into 6 groups (n = 18 each): control, PEMF, FK506, PEMF + hDPSCs, PEMF + FK506, and PEMF + hDPSCs + FK506 groups. hDPSCs (cell = 1 × 106/10 μl/rat) were injected at the crush site immediate after injury. FK506 was administered 3 weeks in FK506 group (0.5 mg/kg/d) while pre-op 1 d and post-op 7 d in PEMF + FK506 and PEMF + hDPSCs + FK506 group; cell tracking was done with PKH26-labeled hDPSCs (cell = 1 × 106/10 μl/rat). The rats were follow-up for 3 weeks. Results. PEMF + FK506 and PEMF + hDPSCs + FK506 group showed a sharp increase in sciatic function index (SFI), axon counts, densities, and labeled neurons in dorsal root ganglia (DRG) than control at 3 weeks. Other three treatment groups also showed higher axon counts, densities, and labeled neurons than control. Higher axon counts and densities were found in PEMF + FK506 and PEMF + hDPSCs + FK506 groups comparing with PEMF group. Brain-derived neurotrophic factor (BDNF) mRNA expression pattern in nerve segment and DRG was almost same. Higher expression level in all the treatment groups was discovered in the follow-up period, but there was no significant difference. Conclusions. All treatment groups can improve regeneration of neurons following crushed injury, PEMF + FK506 and PEMF + hDPSCs + FK506 groups showed higher regeneration ability than other three groups. FK506 plays an important role during hDPSCs transplantation.

Simultaneous Inferior Alveolar Nerve Regeneration and Osseointegration With a Nerve Growth Factor-Supplying Implant: A Preliminary Study

PURPOSE Although nerve growth factor (NGF) has been proved to enhance inferior alveolar nerve (IAN) regeneration, its clinical application remains a challenging issue. This study investigated the functional regeneration of IAN injury by supplying NGF using an NGF-supplying implant and its effect on the osseointegration. MATERIALS AND METHODS In canine IAN transection-and-repair models (n = 9), NGF-supplying implants connected to osmotic pumps were installed just above the transection site. In the right IAN, NGF 300 μg in phosphate buffered saline (PBS) 2 mL was loaded in the pump and pure PBS 2 mL was loaded in the left IAN. The gross clinical finding was evaluated by wound healing, inflammation, implant exposure, and loss of fixture. To evaluate IAN regeneration, electrophysiologic (amplitude, latency, conduction velocity, and peak voltage) and histomorphometric (axon count and density, myelin thickness, and ratio of axon diameter to fiber diameter) analyses were performed. Implant stability quotient, bone-to-implant contact ratio, and new bone area were measured to assess the osseointegration of the NGF-supplying implant. RESULTS The conduction velocity (2.675 m/second) and peak voltage (1.940 μV) of the NGF group at 6 weeks were considerably higher than those of the PBS group (1.892 m/second and 1.300 μV, respectively). The same results were observed for axon count (NGF vs PBS, 4,576.107 ± 270.413 vs 3,606.972 ± 242.876), axon density (10,707.458 ± 638.835 vs 7,899.781 ± 1,063.625/mm(2)), and myelin thickness (1.670 ± 0.555 vs 1.173 ± 0.388 μm). There were no meaningful differences for the other parameters. CONCLUSIONS Supplying NGF with specially designed dental implants can be a new therapeutic approach to enable IAN regeneration and osseointegration simultaneously.

Increased depression-like behaviors with dysfunctions in the stress axis and the reward center by free access to highly palatable food

This study was conducted to examine the behavioral consequences of unlimited consumption of highly palatable food (HPF) and investigate its underlying neural mechanisms. Male Sprague-Dawley rats had free access to chocolate cookie rich in fat (HPF) in addition to ad libitum chow and the control group received chow only. Rats were subjected to behavioral tests during the 2nd week of food condition; i.e. ambulatory activity test on the 8th, elevated plus maze test (EPM) on the 10th and forced swim test (FST) on the 14th day of food condition. After 8 days of food condition, another group of rats were placed in a restraint box and tail bloods were collected at 0, 20, 60, and 120 time points during 2h of restraint period, used for the plasma corticosterone assay. At the end of restraint session, rats were sacrificed and the tissue sections of the nucleus accumbens (NAc) were processed for c-Fos immunohistochemistry. Ambulatory activities and the scores of EPM were not significantly affected by unlimited cookie consumption. However, immobility duration during FST was increased, and swim decreased, in the rats received free cookie access compared with control rats. Stress-induced corticosterone increase was exaggerated in cookie-fed rats, while the stress-induced c-Fos expression in the NAc was blunted, compared to control rats. Results suggest that free access to HPF may lead to the development of depression-like behaviors in rats, likely in relation with dysfunctions in the hypothalamic-pituitary-adrenal axis and the reward center.

Low-frequency pulsed electromagnetic field pretreated bone marrow-derived mesenchymal stem cells promote the regeneration of crush-injured rat mental nerve

Bone marrow-derived mesenchymal stem cells (BMSCs) have been shown to promote the regeneration of injured peripheral nerves. Pulsed electromagnetic field (PEMF) reportedly promotes the proliferation and neuronal differentiation of BMSCs. Low-frequency PEMF can induce the neuronal differentiation of BMSCs in the absence of nerve growth factors. This study was designed to investigate the effects of low-frequency PEMF pretreatment on the proliferation and function of BMSCs and the effects of low-frequency PEMF pre-treated BMSCs on the regeneration of injured peripheral nerve using in vitro and in vivo experiments. In in vitro experiments, quantitative DNA analysis was performed to determine the proliferation of BMSCs, and reverse transcription-polymerase chain reaction was performed to detect S100 (Schwann cell marker), glial fibrillary acidic protein (astrocyte marker), and brain-derived neurotrophic factor and nerve growth factor (neurotrophic factors) mRNA expression. In the in vivo experiments, rat models of crush-injured mental nerve established using clamp method were randomly injected with low-frequency PEMF pretreated BMSCs, unpretreated BMSCs or PBS at the injury site (1 × 106 cells). DiI-labeled BMSCs injected at the injury site were counted under the fluorescence microscope to determine cell survival. One or two weeks after cell injection, functional recovery of the injured nerve was assessed using the sensory test with von Frey filaments. Two weeks after cell injection, axonal regeneration was evaluated using histomorphometric analysis and retrograde labeling of trigeminal ganglion neurons. In vitro experiment results revealed that low-frequency PEMF pretreated BMSCs proliferated faster and had greater mRNA expression of growth factors than unpretreated BMSCs. In vivo experiment results revealed that compared with injection of unpretreated BMSCs, injection of low-frequency PEMF pretreated BMSCs led to higher myelinated axon count and axon density and more DiI-labeled neurons in the trigeminal ganglia, contributing to rapider functional recovery of injured mental nerve. These findings suggest that low-frequency PEMF pretreatment is a promising approach to enhance the efficacy of cell therapy for peripheral nerve injury repair.

Early life stress experience may blunt hypothalamic leptin signalling

The aim of this study was to investigate whether neonatal maternal separation (MS) – chronic stress experience in early life – affects the anorectic efficacy of leptin in the offspring at adolescence. Sprague–Dawley pups were separated from the dam daily for 3 h during postnatal day 1–14 or left undisturbed as non-handled controls (NH). NH and MS male pups received an intraperitoneal leptin (100 μg/kg) or saline on postnatal day (PND) 28, and then food intake and body weight gain were recorded. The hypothalamic levels of leptin-signalling-related genes, phosphorylated signal transducer and activator of transcription-3 (pSTAT3) and protein-tyrosine phosphatase 1B (PTP1B) were examined at 40 min after a single injection of leptin on PND 39 by immunohistochemistry and Western blot analysis. Leptin-induced suppressions in food intake and weight gain was observed in NH pups, but not in MS. Leptin increased pSTAT3 in the hypothalamic arcuate nucleus of NH pups, but not of MS. Interestingly, basal levels of the hypothalamic PTP1B and pSTAT3 were increased in MS pups compared with NH controls. The results suggest that neonatal MS experience may blunt the anorectic efficacy of leptin later in life, possibly in relation with increased expressions of PTP1B and/or pSTAT3 in the hypothalamus.

Isolation and Purification of Schwann Cells from Human Peripheral Nerves via the Cold Jet Technique

Schwann cells (SCs) play an important role in peripheral nerve regeneration. However, established methods for SC preparation have limitations in that they are non-specific and time-consuming. The purpose of this study was to assess the amount and purity of SCs isolated from human peripheral nerve tissue using the cold jet technique. The cold jet technique is an SC isolation and purification method that exploits the different responses of the SCs and fibroblasts to temperature shock. Specifically, the temperature shock causes specific detachment of SCs from fibroblasts. A total of 32 human peripheral nerve tissues (sensory and motor) were obtained from reconstruction procedures for oral cancer. After the length and weight of nerve segments were measured, the nerve fascicles were digested and SCs were isolated and purified by the cold jet technique. The purity of SCs was confirmed by morphology, RT-PCR, immunohistochemistry and flow cytometry. Phasecontrast photomicrographs, S100 mRNA expression rates and immunohistochemical staining show that the SC purification rate was greater with the cold jet technique. Flow cytometry showed the percentage (%) gated value increased from 56.10 to 95.95%. On average, 2.48 ± 2.57 × 106 SCs/g or 0.16 ± 0.18 × 106 SCs/cm were harvested efficiently from human peripheral nerve segments. The cold jet technique was an efficient method for SC isolation and purification from the human peripheral nerves. The determined amount and the level of purity of the SC harvested by this method may be helpful for tissue engineering or cell therapy