Hyaluronic Acid Conjugated with 17ß-Estradiol Effectively Alleviates Estropause-Induced Cognitive Deficits in Rats.

Women are at a higher risk of cognitive impairments and Alzheimer's disease (AD), particularly after the menopause, when the estrous cycle becomes irregular and diminishes. Numerous studies have shown that estrogen deficiency, especially estradiol (E2) deficiency, plays a key role in this phenomenon. Recently, a novel polymeric drug, hyaluronic acid-17ß-estradiol conjugate (HA-E2), has been introduced for the delivery of E2 to brain tissues. Studies have indicated that HA-E2 crosses the blood-brain barrier (BBB) and facilitates a prolonged E2 release profile while lowering the risk of estrogen-supplement-related side effects. In this study, we used ovariohysterectomy (OHE) rats, a postmenopausal cognitive deficit model, to explore the effect of a 2-week HA-E2 treatment (210 ng/kg body weight, twice a week) on the cholinergic septo-hippocampal innervation system, synaptic transmission in hippocampal pyramidal neurons and cognitive improvements. Our study revealed an 11% rise in choline acetyltransferase (ChAT) expression in both the medial septal nucleus (MS nucleus) and the hippocampus, along with a 14-18% increase in dendritic spine density in hippocampal pyramidal neurons, following HA-E2 treatment in OHE rats. These enhancements prompted the recovery of cognitive functions such as spatial learning and memory. These findings suggest that HA-E2 may prevent and improve estrogen-deficiency-induced cognitive impairment and AD.

The effect of astaxanthin treatment on the rat model of fetal alcohol spectrum disorders (FASD).

Fetal alcohol spectrum disorder (FASD) caused by mother's exposure to alcohol during pregnancy is a congenital neurological disease of the fetus resulting in fetal developmental and intellectual disabilities, cognitive impairment, and coordination disorder. Excess oxidative stress and neuroinflammatory responses were an important factor in neuropathological changes in FASD. Astaxanthin (AST) was a potent antioxidant and anti-inflammatory carotenoid. Therefore, this study proposed to explore how AST treatment can ameliorate morphological changes in the hippocampus and cognitive impairment in FASD rats by reducing oxidative stress and neuroinflammation in the brain. An alcohol atomizer was used from postnatal day (P) 2 to P10 to induce the FASD rat model. They were treated with AST (10 mg/kg body weight/day, intraperitoneal injection) for 8 consecutive days starting at P53 and sacrificed at P60. FASD rats had growth retardation and facial dysmorphologies, excessive oxidative stress and neuroinflammation in the hippocampus, decreased choline acetyltransferase (ChAT) expression in MS nucleus, spine loss on hippocampal CA1 pyramidal neurons, and poor performance in spatial learning and memory and sensory-motor coordination. After AST treatment, oxidative stress, neuroinflammation, cholinergic system, excitatory synaptic structure and behavior of FASD rats improved. Therefore, our study provided evidence to support the proposal that AST could be considered to treat FASD.

The effects of astaxanthin treatment on a rat model of Alzheimer's disease.

Alzheimer's disease (AD), a progressive neurodegenerative disorder characterized by memory loss and dementia, could be a consequence of the abnormalities of cortical milieu, such as oxidative stress, inflammation, and/or accompanied with the aggregation of ß-amyloid. The majority of AD patients are sporadic, late-onset AD, which predominantly occurs over 65 years of age. Our results revealed that the ferrous amyloid buthionine (FAB)-infused sporadic AD-like model showed deficits in spatial learning and memory and with apparent loss of choline acetyltransferase (ChAT) expression in medial septal (MS) nucleus. In hippocampal CA1 region, the loss of pyramidal neurons was accompanied with cholinergic fiber loss and neuroinflammatory responses including glial reaction and enhanced expression of inducible nitric oxide synthase (iNOS). Surviving hippocampal CA1 pyramidal neurons showed the reduction of dendritic spines as well. Astaxanthin (ATX), a potent antioxidant, reported to improve the outcome of oxidative-stress-related diseases. The ATX treatment in FAB-infused rats decreased neuroinflammation and restored the ChAT + fibers in hippocampal CA1 region and the ChAT expression in MS nucleus. It also partly recovered the spine loss on hippocampal CA1 pyramidal neurons and ameliorated the behavioral deficits in AD-like rats. From these data, we believed that the ATX can be a potential option for slowing the progression of Alzheimer's disease.

Reproductive experience modified dendritic spines on cortical pyramidal neurons to enhance sensory perception and spatial learning in rats.

Behavioral adaptations during motherhood are aimed at increasing reproductive success. Alterations of hormones during motherhood could trigger brain morphological changes to underlie behavioral alterations. Here we investigated whether motherhood changes a rat's sensory perception and spatial memory in conjunction with cortical neuronal structural changes. Female rats of different statuses, including virgin, pregnant, lactating, and primiparous rats were studied. Behavioral test showed that the lactating rats were most sensitive to heat, while rats with motherhood and reproduction experience outperformed virgin rats in a water maze task. By intracellular dye injection and computer-assisted 3-dimensional reconstruction, the dendritic arbors and spines of the layer III and V pyramidal neurons of the somatosensory cortex and CA1 hippocampal pyramidal neurons were revealed for closer analysis. The results showed that motherhood and reproductive experience increased dendritic spines but not arbors or the lengths of the layer III and V pyramidal neurons of the somatosensory cortex and CA1 hippocampal pyramidal neurons. In addition, lactating rats had a higher incidence of spines than pregnant or primiparous rats. The increase of dendritic spines was coupled with increased expression of the glutamatergic postsynaptic marker protein (PSD-95), especially in lactating rats. On the basis of the present results, it is concluded that motherhood enhanced rat sensory perception and spatial memory and was accompanied by increases in dendritic spines on output neurons of the somatosensory cortex and CA1 hippocampus. The effect was sustained for at least 6 weeks after the weaning of the pups.

Exogenous dehydroisoandrosterone sulfate reverses the dendritic changes of the central neurons in aging male rats.

Sex hormones are known to help maintaining the cognitive ability in male and female rats. Hypogonadism results in the reduction of the dendritic spines of central neurons which is believed to undermine memory and cognition and cause fatigue and poor concentration. In our previous studies, we have reported age-related regression in dendrite arbors along with loss of dendritic spines in the primary somatosensory cortical neurons in female rats. Furthermore, castration caused a reduction of dendritic spines in adult male rats. In light of this, it was surmised that dendritic structures might change in normal aging male rats with advancing age. Recently, dehydroepiandrosterone sulfate (DHEAS) has been reported to have memory-enhancing properties in aged rodents. In this study, normal aging male rats, with a reduced plasma testosterone level of 75-80%, were used to explore the changes in behavioral performance of neuronal dendritic arbor and spine density. Aging rats performed poorer in spatial learning memory (Morris water maze). Concomitantly, these rats showed regressed dendritic arbors and spine loss on the primary somatosensory cortical and hippocampal CA1 pyramidal neurons. Exogenous DHEAS and testosterone treatment reversed the behavioral deficits and partially restored the spine loss of cortical neurons in aging male rats but had no effects on the dendritic arbor shrinkage of the affected neurons. It is concluded therefore that DHEAS, has the efficacy as testosterone, and that it can exert its effects on the central neuron level to effectively ameliorate aging symptoms.

Genistein partly eases aging and estropause-induced primary cortical neuronal changes in rats.

Gonadal hormones can modulate brain morphology and behavior. Recent studies have shown that hypogonadism could result in cortical function deficits. To this end, hormone therapy has been used to ease associated symptoms but the risk may outweigh the benefits. Here we explored whether genistein, a phytoestrogen, is effective in restoring the cognitive and central neuronal changes in late middle age and surgically estropause female rats. Both animal groups showed poorer spatial learning than young adults. The dendritic arbors and spines of the somatosensory cortical and CA1 hippocampal pyramidal neurons were revealed with intracellular dye injection and analyzed. The results showed that dendritic spines on these neurons were significantly decreased. Remarkably, genistein treatment rescued spatial learning deficits and restored the spine density on all neurons in the surgically estropause young females. In late middle age females, genistein was as effective as estradiol in restoring spines; however, the recovery was less thorough than on young OHE rats. Neither genistein nor estradiol rectified the shortened dendritic arbors of the aging cortical pyramidal neurons suggesting that dendritic arbors and spines are differently modulated. Thus, genistein could work at central level to restore excitatory connectivity and appears to be potent alternative to estradiol for easing aging and menopausal syndromes.

Morphological changes of cortical pyramidal neurons in hepatic encephalopathy.

BACKGROUND: Hepatic encephalopathy (HE) is a reversible neuropsychiatric syndrome associated with acute and chronic liver diseases. It includes a number of neuropsychiatric disturbances including impaired motor activity and coordination, intellectual and cognitive function. RESULTS: In the present study, we used a chronic rat HE model by ligation of the bile duct (BDL) for 4 weeks. These rats showed increased plasma ammonia level, bile duct hyperplasia and impaired spatial learning memory and motor coordination when tested with Rota-rod and Morris water maze tests, respectively. By immunohistochemistry, the cerebral cortex showed swelling of astrocytes and microglia activation. To gain a better understanding of the effect of HE on the brain, the dendritic arbors of layer V cortical pyramidal neurons and hippocampal CA1 pyramidal neurons were revealed by an intracellular dye injection combined with a 3-dimensional reconstruction. Although the dendritic arbors remained unaltered, the dendritic spine density on these neurons was significantly reduced. It was suggested that the reduction of dendritic spines may be the underlying cause for increased motor evoked potential threshold and prolonged central motor conduction time in clinical finding in cirrhosis. CONCLUSIONS: We found that HE perturbs CNS functions by altering the dendritic morphology of cortical and hippocampal pyramidal neurons, which may be the underlying cause for the motor and intellectual impairments associated with HE patients.

The distribution of muscles fibers and their types in the female rat urethra: cytoarchitecture and three-dimensional reconstruction.

An attempt to explore urethral cytoarchitecture including the distribution of smooth muscles and fast and slow striated muscles of adult female Sprague Dawley rat--a popular model in studying lower urinary tract function. Histological and immunohistochemical stainings were carried out to investigate the distribution of urethral muscle fibers and motor end plates. The urethral sphincter was furthermore three-dimensionally reconstructed from serial histological sections. The mucosa at the distal urethra was significantly thicker than that of other segments. A prominent inner longitudinal and outer circular layer of smooth muscles covered the proximal end of urethra. Thick circular smooth muscles of the bladder neck region (urethral portion) decreased significantly distalward and longitudinal smooth muscles became 2- to 3-fold thicker in the rest of the urethra. An additional layer of striated muscles appeared externally after neck region (urethra) and in association with motor end plates ran throughout the remaining urethra as the striated sphincter layer. Most striated muscles were fast fibers while relatively fewer slow fibers often concentrated at the periphery. A pair of extraneous striated muscles, resembling the human urethrovaginal sphincter muscles, connected both sides of mainly the distal vagina to the dorsal striated muscles in the wall of the middle urethra. The tension provided by this pair of muscles, and in conjunction with the striated sphincter of the urethral wall, was likely to function to suspend the middle urethra and facilitates its closure. Comprehensive morphological data of urethral sphincter offers solid basis for researchers conducting studies on dysfunction of bladder outlet.

Testosterone modulation of dendritic spines of somatosensory cortical pyramidal neurons.

Brain structures and functions are increasingly recognized to be directly affected by gonadal hormones, which classically determine reproductive functions and sexual phenotypes. In this regard, we found recently that ovariectomy trimmed the dendritic spines of female rat primary somatosensory cortical neurons and estradiol supplement reversed it. Here, we investigated whether in the male androgen also has a cortical modulatory effect. The dendritic arbors and spines of rat somatosensory cortical pyramidal neurons were studied following intracellular dye injection and three-dimensional reconstruction. Dendritic spines, but not length, of the layers III and V pyramidal neurons were found reduced at 2 weeks and rebounded slightly at 4 weeks and further at 8 and 24 weeks following castration, which, however, remained significantly fewer than those of the intact animals. Two weeks of osmotic pump-delivered testosterone treatment to animals castrated for 4 weeks replenished serum testosterone and reversed the densities of dendritic spines on these neurons to control animal levels. Androgen receptor appears to mediate this effect as its antagonist flutamide reduced the dendritic spines of normal adult rats while causing a mild feedback surge of serum testosterone. On the other hand, blocking the conversion of testosterone to estrogen with the aromatase inhibitor anastrozole failed to alter the dendritic spine densities in male adult rats. In conclusion, these results support our hypothesis that testosterone acts directly on the androgen receptor in males to modulate the dendritic spines of somatosensory cortical output neurons.

Gonadal hormones modulate the dendritic spine densities of primary cortical pyramidal neurons in adult female rat.

Adult dendritic arbors and spines can be modulated by environment and gonadal hormones that have been reported to affect also those of hippocampal and prefrontal cortical neurons. Here we investigated whether female gonadal hormones and estrous cycle alter the dendrites of primary cortical neurons. We employed intracellular dye injection in semifixed brain slices and 3-dimensional reconstruction to study the dendritic arbors and spines of the major cortical output cells, layer III and V pyramidal neurons, during different stages of the estrous cycle. Dendritic spines of both pyramidal neurons were more numerous during proestrus than estrus and diestrus, whereas dendritic arbors remained unaffected. Ovariohysterectomy (OHE) reduced dendritic spines by 24-30% in 2 weeks, whereas subcutaneous estrogen or progesterone supplement restored it to normal estrous/diestrous level in 14 days; neither treatment affected the dendritic arbors. Reduction of dendritic spines following OHE was associated with decrease of PSD-95 suggesting decrease of excitatory synapses. Thus, fluctuation of gonadal hormones during the female sex cycle is likely to modulate primary cortical functions and loss of gonadal hormones for instance following menopause might compromise cortical function, and the effect could be reversed by exogenous female sex hormones.

Transplanted bone marrow stromal cells migrate, differentiate and improve motor function in rats with experimentally induced cerebral stroke.

Bone marrow stromal cells are multipotential cells that can be induced to differentiate into osteoblasts, chondrocytes, myocytes and adipocytes in different microenvironments. Recent studies revealed that bone marrow stromal cells could improve neurological deficits of various damages or diseases of the central nervous system such as Parkinson's disease, brain trauma, spinal cord injury and multiple sclerosis, and promote glia-axonal remodeling in animal brain subjected to an experimentally induced stroke. In the present study, bone marrow stromal cells were intracerebrally transplanted into the cerebrum following a transient middle cerebral artery occlusion. Our aim was to find out whether the bone marrow stromal cells could survive and express neural phenotypic proteins and, in addition, whether they could restore the behavioral and functional deficits of the cerebral ischemic rats. Our results demonstrated that transplanted bone marrow stromal cells survived and migrated to areas around the lesion site. Some of them exhibited marker proteins of astrocytes and oligodendrocytes. Bone marrow stromal cell implantation significantly reduced the transient middle cerebral artery occlusion-induced cortical loss and thinning of the white matter and enhanced cortical beta-III-tubulin immunoreactivity. Rats implanted with bone marrow stromal cells showed significant improvement in their performance of elevated body swing test and forelimb footprint analysis and only transient recovery of the adhesive-removal test. Our data support bone marrow stromal cells as a valuable source of autologous or allogenic donor cells for transplantation to improve the outcome following cerebral ischemia.

Neurogenesis of cuneothalamic neurons and NO-containing neurons in the cuneate nucleus of the rat.

The genesis of the cuneothalamic neurons (CTNs) in the rat cuneate nucleus was determined by a double-labeling method using 5'-bromodeoxyuridine (BrdU), the thymidine analogue, and Fluoro-Gold (FG), a retrograde fluorescent tracer. BrdU-positive cells were observed in the cuneate nucleus in all rats receiving BrdU injection at embryonic days (E) E13--E16; none was detected in rats given BrdU injection at E12. At E13 and E14, BrdU-positive cells were randomly distributed. However, at E15, the number of BrdU-positive cells was clearly reduced and the majority of them was located at the dorsolateral or peripheral region of the nucleus. FG/BrdU double-labeling study showed the existence of BrdU-labeled CTNs when the mother rat received BrdU injection at E13 and E14, being more numerous at E13 in which the neurons were scattered throughout the nucleus. At E14, however, the majority of the BrdU-labeled CTNs were located superficially in the nucleus. Double-labeled cells were undetected in rats that had been exposed to BrdU at E15 and E16. Quantitative data showed that the majority (ca 70-80%) of the CTNs were generated at E13, and were markedly decreased at E14 (ca 4-6%). Using nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry coupled with BrdU immunohistochemistry, we have shown the NADPH-d/BrdU double-labeled neurons in the nucleus between E13 and E15, with the majority of them occurring at E14, but absent at E16. The present results suggest that the CTNs are generated prior to the NO-containing neurons in the cuneate nucleus.