Polypide anatomy of hornerid bryozoans (Stenolaemata: Cyclostomatida).

Bryozoans are small colonial coelomates whose colonies are made of individual modules (zooids). Like most coelomate animals, bryozoans have a characteristic body wall composition, including an epidermis, an extracellular matrix (ECM) and a coelothelium, all pressed together. The order Cyclostomatida, however, presents the most striking deviation, in which the ECM and the corresponding coelothelium underlying major parts of the skeletal wall epidermis are detached to form an independent membranous sac. It forms a separate, much smaller compartment, suspended in the zooid body cavity and working as an important element of the cyclostome lophophore protrusion mechanism. The polypide anatomy and ultrastructure of this group is best known from studies of one family, the Crisiidae (Articulata). Here, we examined four species from the phylogenetically and ecologically contrasting family Horneridae (Cancellata) from New Zealand, and provide the first detailed ultrastructural description of the hornerid polypide, including tentacles, mouth region, digestive system and the funiculus. We were able to trace continuity and transitions of cell and ECM layers throughout the whole polypide. In addition, we identified that the funiculus is a lumen-free ECM cord with two associated muscles, disconnected from interzooidal pores. Except for funicular core composition, the polypide anatomy of hornerids agrees well with the general cyclostomate body plan.

Single neuron activity and c-Fos expression in the rat striatum following electrical stimulation of the peripheral vestibular system.

Connections between the vestibular system and the basal ganglia have been postulated since the early 20th century. However, the results of electrophysiological studies investigating neuronal responses to electrical stimulation of the vestibular system have been inconsistent. The aim of this study was to investigate the effects of electrical stimulation of the vestibular labyrinth on single neuron activity and c-Fos expression in the rat striatum. We used electrical stimulation of the vestibular labyrinth (various intensities delivered to the round window) to examine the electrophysiological response of striatal neurons and c-Fos expression. From 507 single neurons recorded (n = 20 rats), no vestibular-responsive neuron was found at 1× and 2× the nystagmus threshold; however, 6 neurons were found at 3× the threshold. These neurons were found bilaterally, with a response latency of ~50 msec from the end of the stimulus. For the c-Fos study, the number of neurons expressing c-Fos was quantified using stereological methods. Stimulation at 2× the threshold for nystagmus (n = 5 rats) resulted in a significant decrease in the number of neurons expressing c-Fos in the bilateral striatum compared to both the sham control group (n = 5) and the lower stimulus intensity group (n = 5) (P ≤ 0.0001 for both). The results of this study demonstrate that: (1) some single striatal neurons respond to electrical vestibular stimulation, however, these responses are circumscribed and infrequent; (2) electrical stimulation of the vestibular labyrinth results in a decrease in the number of striatal neurons expressing c-Fos, in a current-dependent manner.

Total Number Is Important: Using the Disector Method in Design-Based Stereology to Understand the Structure of the Rodent Brain.

The advantages of using design-based stereology in the collection of quantitative data, have been highlighted, in numerous publications, since the description of the disector method by Sterio (1984). This review article discusses the importance of total number derived with the disector method, as a key variable that must continue to be used to understand the rodent brain and that such data can be used to develop quantitative networks of the brain. The review article will highlight the huge impact total number has had on our understanding of the rodent brain and it will suggest that neuroscientists need to be aware of the increasing number of studies where density, not total number, is the quantitative measure used. It will emphasize that density can result in data that is misleading, most often in an unknown direction, and that we run the risk of this type of data being accepted into the collective neuroscience knowledge database. It will also suggest that design-based stereology using the disector method, can be used alongside recent developments in electron microscopy, such as serial block-face scanning electron microscopy (SEM), to obtain total number data very efficiently at the ultrastructural level. Throughout the article total number is discussed as a key parameter in understanding the micro-networks of the rodent brain as they can be represented as both anatomical and quantitative networks.

Basal dendritic length is reduced in the rat hippocampus following bilateral vestibular deafferentation.

Some previous studies in humans have shown that bilateral loss of vestibular function is associated with a significant bilateral atrophy of the hippocampus, which correlated with the patients' spatial memory deficits. By contrast, studies in rats have failed to detect any changes in hippocampal volume following bilateral vestibular loss. Therefore, in this study we investigated whether bilateral vestibular deafferentation (BVD) might result in more subtle morphological changes in the rat hippocampus, involving alterations in dendritic intersections, using Golgi staining and Sholl analysis. We found that at 1month following BVD, there was a significant decrease in basal (P⩽0.0001) but not apical dendritic intersections in the CA1 region of the hippocampus compared to sham-operated animals and anaesthetic controls. However, dendritic branching was not significantly affected. These results suggest that the rat hippocampus does undergo subtle morphological changes following bilateral vestibular loss, and that they may be in the form of alterations in dendritic structure.

Apoptotic cell death and temporal expression of apoptotic proteins Bcl-2 and Bax in the hippocampus, following binge ethanol in the neonatal rat model.

BACKGROUND: Binge-like ethanol (EtOH) exposure during the early rat neonatal period results in acute cell loss in specific brain regions, but such acute cell death has not been well established in the hippocampus. Binge alcohol exposure can also result in protein expression changes in the cerebellum that could alter cell fate, but this has not been reported for the hippocampal subregions. This study investigates acute apoptotic cell death in hippocampal regions CA1, CA3, and dentate gyrus (DG) following a binge EtOH exposure on postnatal day (PN) 6, PN8, or PN6 + 8 and the alteration in pro- and anti-apoptotic proteins following a single EtOH binge on PN6. METHODS: Apoptotic cell death was quantified 12 hours after EtOH binge exposure using the optical fractionator method. Western blot analysis determined expression of pro-apoptotic Bax and anti-apoptotic Bcl-2, 12, 24, and 48 hours after binge EtOH exposure on PN6. The Bcl-2:Bax ratio was used as a measure of vulnerability to apoptosis. RESULTS: Acute apoptosis increased significantly 12 hours following PN6 or 8 EtOH exposure in CA1, CA3, and DG, but the magnitude of apoptotic cell death was significantly greater in CA1 than in CA3 and DG, which did not differ. Significant cell death was not detected when a PN8 EtOH exposure was preceded by exposure on PN6. Binge EtOH exposure on PN6 resulted in a significant increase in expression of Bcl-2 and the Bcl-2:Bax ratio in the CA1/DG region at 24 hours after EtOH exposure on PN6. The Bcl-2:Bax ratio in the CA3 region was not altered. CONCLUSIONS: This study shows that repeated binge exposure does not have a cumulative effect on the magnitude of acute apoptotic cell death. This finding may be explained in part by changes in the Bcl-2:Bax ratio after a single binge EtOH exposure.

The effects of bilateral vestibular loss on hippocampal volume, neuronal number, and cell proliferation in rats.

Previous studies in humans have shown that bilateral loss of vestibular function is associated with a significant bilateral atrophy of the hippocampus, which correlated with the patients' spatial memory deficits. More recently, patients who had recovered from unilateral vestibular neuritis have been reported to exhibit a significant atrophy of the left posterior hippocampus. Therefore, we investigated whether bilateral vestibular deafferentation (BVD) would result in a decrease in neuronal number or volume in the rat hippocampus, using stereological methods. At 16 months post-BVD, we found no significant differences in hippocampal neuronal number or volume compared to sham controls, despite the fact that these animals exhibited severe spatial memory deficits. By contrast, using bromodeoxyuridine (BrdU) as a marker of cell proliferation, we found that the number of BrdU-labeled cells significantly increased in the dentate gyrus of the hippocampus between 48 h and 1 week following BVD. Although a substantial proportion of these cells survived for up to 1 month, the survival rate was significantly lower in BVD animals when compared with that in sham animals. These results suggest a dissociation between the effects of BVD on spatial memory and hippocampal structure in rats and humans, which cannot be explained by an injury-induced increase in cell proliferation.

Acute and long-term Purkinje cell loss following a single ethanol binge during the early third trimester equivalent in the rat.

BACKGROUND: In the rat, binge-like ethanol (EtOH) exposure during the early neonatal period (a developmental period equivalent to the human third trimester) can result in a permanent deficit of cerebellar Purkinje cells (Pcells). However, the consequences of a moderate binge alcohol exposure on a single day during this postnatal period have not been established. This is an issue of importance as many pregnant women binge drink periodically at social drinking levels. This study aimed to identify both the acute and long-term effects of exposure to a single alcohol binge that achieved a mean peak blood EtOH concentration of approximately 250 mg/dl during early postnatal life using a rat model of fetal alcohol spectrum disorders. METHODS: Acute apoptotic Pcell death 10 hours after a moderate dose binge EtOH exposure from postnatal days (PDs) 0 to 10 was assessed using active caspase-3 immunolabeling. Acute Pcell apoptosis was quantified in cerebellar vermal lobules I-X using the physical disector method. Long-term effects were assessed at PD 60 using stereological methods to determine total Pcell numbers in the vermis, lobule III, and lobule IX, following a moderate dose binge EtOH exposure at PDs 0, 2, or 4. RESULTS: Acute apoptosis was induced by EtOH on PDs 1 to 8 in a time and lobular-dependent manner. For EtOH exposure on PD 2, significant long-term Pcell loss occurred in lobule III. EtOH exposure on PD 4 resulted in significant long-term Pcell loss throughout the entire vermis. CONCLUSIONS: These results indicate that a single, early EtOH episode of moderate dose can create significant and permanent Pcell loss in the developing cerebellum.

Early postnatal exposure to alcohol reduces the number of neurons in the occipital but not the parietal cortex of the rat.

BACKGROUND: The rat brain undergoes a period of rapid growth in the early postnatal period. During this time, the neocortex seems to be vulnerable to ethanol injury. Subdivisions of the neocortex develop in a temporospatial gradient that is likely to determine their vulnerability to ethanol-induced damage and whether damage is permanent. Therefore, the authors investigated the effect of postnatal ethanol exposure on the neocortex and specific subregions at the cessation of exposure and in the mature brain. METHODS: Four-day-old rat pups with intragastric cannulae were artificially reared from postnatal day (PN) 4 through PN9. Of 12 daily feeds, two consecutive feeds contained either ethanol (4.5 g/kg) or an isocaloric maltose/dextrin solution. On PN10 or PN115, animals were perfused intracardially, and the brains were removed. Stereological methods were used to determine the total number of neurons and glial cells in, and the volume of, the neocortex, the parietal cortex, and the occipital cortex. RESULTS: Exposure to ethanol did not affect body or brain weight at PN10. In contrast, at PN115 forebrain weight was significantly lower in ethanol-exposed animals compared with control-treated animals. There was no effect of treatment on body weight at PN115. On PN10, neocortical volume was 15% smaller in the ethanol-exposed animals compared with controls, with no change in the total number of neurons or glial cells. Occipital cortical volume was reduced by 22% in the ethanol-exposed animals, with a significant deficit in the total number of neurons (ethanol-exposed, 2.62 x 10; gastrostomy control, 3.20 x 10). There was no effect of ethanol exposure on the total number of glial cells in the occipital cortex or on any parameter in the parietal cortex. There was also no significant effect of ethanol exposure on the occipital cortex on PN115. CONCLUSIONS: These findings provide support for the hypothesis that a specific area or cell population might be differentially vulnerable to ethanol exposure during the brain growth spurt and that cell deficits evident on PN10 may not be permanent.

Foraging experience, glomerulus volume, and synapse number: A stereological study of the honey bee antennal lobe.

The primary antennal sensory centers (antennal lobes) in the brain of the honeybee are highly compartmentalized into discrete spheres of synaptic neuropil called glomeruli. Many of the glomeruli can be identified according to their predictable size and location. This study examines T1-44, a prominent glomerulus on the dorsal surface of the antennal lobe. Previously, we have shown that the volume of T1-44 in 4-day-old workers performing tasks within the hive is significantly smaller than in foragers and that increases in volume are accompanied by an increase in total synapse number in this glomerulus. Here we examine whether foraging experience is essential for either changes in volume or for changes in synapse numbers in glomerulus T1-44. Five-day-old bees reared under normal colony conditions were compared with 5-day-old bees reared under isolated conditions, and also to 5-day-old bees that had been induced to forage precociously. A combination of light and electron microscopy was used to compare T1-44 volumes and synapse numbers in these three groups. Two groups of 11-day-old bees, precocious foragers and nonforagers, were also examined. The Cavalieri direct estimator of volume was applied to 1.5 microm sections of resin embedded brains. Selected sections were then re-embedded and prepared for transmission electron microscopy. Synapse densities were determined using the physical disector method on electron micrographs. Synapse density and glomerulus volume were combined to give an unbiased estimate of the total number of synapses. This study shows that while both volume and synapse numbers can be induced to increase prematurely in young (5-day-old) precocious foragers, foraging experience is not essential for these structural changes to occur in glomerulus T1-44.

Stereological analysis reveals striking differences in the structural plasticity of two readily identifiable glomeruli in the antennal lobes of the adult worker honeybee.

The primary antennal sensory centers (antennal lobes) in the brain of the honeybee are highly compartmentalized into discrete spheres of synaptic neuropil called glomeruli, many of which can be identified according to their predictable size and location. Glomeruli undergo significant changes in volume during the lifetime of the adult worker bee, at least some of which are activity dependent. This study tests the commonly expressed assumption that increases in neuropil volume are accompanied by an underlying increase in the number of synapses present in the tissue. A combination of light and electron microscopy was used to determine total synapse number within two glomeruli, T1-44 and T4-2(1). The Cavalieri direct estimator of volume was applied to 1.5 microm sections of resin-embedded brains. Selected sections were then re-embedded and prepared for transmission electron microscopy. Synapse densities were determined using the physical disector method on electron micrographs. Synapse density and glomerulus volume were combined to give an unbiased estimate of the total number of synapses. In glomerulus T1-44, a significant increase in volume was accompanied by a significant increase in the total number of synapses. In contrast, synapse counts in T4-2(1) remained unchanged, despite a significant increase in the volume of this glomerulus. These results demonstrate that synapse proliferation in antennal lobes of the adult worker bee is highly site specific. Although volumetric changes and changes in synapse number both contribute to the structural plasticity of the antennal lobes, these two components of plasticity appear to be independent processes.

Therapeutic effects of complex motor training on motor performance deficits induced by neonatal binge-like alcohol exposure in rats: II. A quantitative stereological study of synaptic plasticity in female rat cerebellum.

Twenty days of complex motor skill training in adult rats was previously demonstrated to rehabilitate motor performance deficits induced by binge alcohol exposure in neonatal rats. This follow-up study evaluated morphological plasticity in the paramedian lobule of the cerebellum (PML) using the same treatment and training regimens. On postnatal days (PD) 4-9, female Long-Evans rats were given either alcohol (Alcohol Exposure - AE, 4.5 g/kg/day via artificial rearing), exposure to gastrostomy control (GC) artificial rearing procedures, or reared normally as suckle controls (SC). After weaning, all rats were housed two to three per cage. At 180 days old, rats were randomly assigned either to a rehabilitation condition (RC: given 20 days of complex motor skill training), or to an inactive condition (IC: remained in their home cage). The AE rats were delayed in acquiring the training, but there were no group differences in performance over the last 2 weeks of training. Unbiased stereological techniques were used to evaluate PML volume, Purkinje cell and parallel fiber synapse density. Although total volume of PML was significantly reduced in the AE rats, complex motor skill training resulted in a significant increase in the PML molecular layer in all three postnatal treatment groups. The RC animals from the SC and AE groups had more parallel fiber synapses per Purkinje cell than corresponding IC animals. These data support the hypothesis that 'rehabilitative' motor training stimulates synaptogenesis in the PML, and that Purkinje neurons that survive the early postnatal alcohol insult are capable of substantial experience-induced plasticity.