bluuebunny/crossref_metadata_2025 huggingface dataset:
Undergraduate ophthalmology education survey of New Zealand ophthalmologists, general practitioners and optometrists
Authors: Ah‐Chan John J , Sanderson Gordon , Vote Brendan JT , Molteno Anthony CB
Published: September 2003

Aim: To determine what New Zealand ophthalmologists, general practitioners and optometrists consider important ophthalmic topic areas requiring emphasis in the medical undergraduate curriculum.

Method: A total of 793 questionnaires related to the content and teaching of undergraduate ophthalmology were sent to ophthalmologists, general practitioners and optometrists. Results were analysed separately for the three respondent groups and as a whole.

Results: Four hundred and fourteen questionnaires were returned (52% return rate). Overall responses of the three participant groups were similar and agreed favourably with the current curriculum. The ability to measure visual acuity (97%) and pupillary reflexes (93%), perform ophthalmoscopy (92%), and assess visual fields (68%) were regarded as ‘important or essential’ by the majority of respondents. Only 53% of respondents consider the ability to diagnose chronic open angle glaucoma as important. The respondents stressed the importance of the diagnosis of predominantly anterior segment disease contrasting with the traditional bias towards the teaching of ophthalmoscopy and posterior segment disease. The majority of respondents stressed the importance of graduating medical students being able to treat bacterial and allergic conjunctivitis, styes, blepharitis, corneal abrasion, and corneal and conjunctival foreign bodies, areas present but not normally emphasized in current curricula.

Conclusion: The findings of this study provided additional data to facilitate curriculum design and illustrated the value of an integrated problem‐based learning approach in ophthalmology undergraduate teaching.
Delayed diffuse lamellar keratitis after laser in situ keratomileusis
Authors: Yeoh Jonathan , Moshegov Con N
Published: September 2003

Two cases are reported of delayed diffuse lamellar keratitis after uneventful laser in situ keratomileusis. The first patient presented with an epithelial defect 6 weeks after laser in situ keratomileusis. Three days later the defect was healed but diffuse lamellar keratitis was noted. This was treated with topical dexamethasone and ketorolac with complete resolution of the diffuse lamellar keratitis over 3 weeks. The second patient presented with an epithelial defect and gross diffuse lamellar keratitis 10 weeks after laser in situ keratomileusis. Treatment was with topical dexamethasone and ciprofloxacin with gradual resolution of the diffuse lamellar keratitis. Common to both patients was a background of rosacea, implanted debris with no initial reaction, and epithelial defects leading to diffuse lamellar keratitis. It is suggested that these two cases represent epithelial defect associated corneal infiltration, which resembles classical diffuse lamellar keratitis with the spread of inflammatory cells through a path of least resistance.
Fretting fatigue behaviour of shot‐peened Ti‐6Al‐4V at room and elevated temperatures
Authors: LEE H. , JIN O. , MALL S.
Published: September 2003

Fretting fatigue behaviour of shot‐peened titanium alloy, Ti‐6Al‐4V was investigated at room and elevated temperatures. Constant amplitude fretting fatigue tests were conducted over a wide range of maximum stresses, σmax= 333 to 666 MPa with a stress ratio of R= 0.1. Two infrared heaters, placed at the front and back of specimen, were used to heat and maintain temperature of the gage section of specimen at 260 °C. Residual stress measurements by X‐ray diffraction method before and after fretting test showed that residual compressive stress was relaxed during fretting fatigue. Elevated temperature induced more residual stress relaxation, which, in turn, decreased fretting fatigue life significantly at 260 °C. Finite element analysis (FEA) showed that the longitudinal tensile stress, σxx varied with the depth inside the specimen from contact surface during fretting fatigue and the largest σxx could exist away from the contact surface in a certain situation. A critical plane based fatigue crack initiation model, modified shear stress range parameter (MSSR), was computed from FEA results to characterize fretting fatigue crack initiation behaviour. It showed that stress relaxation during test affected fretting fatigue life and location of crack initiation significantly. MSSR parameter also predicted crack initiation location, which matched with experimental observations and the number of cycles for crack initiation, which showed the appropriate trend with the experimental observations at both temperatures.
Structural integrity assessment of FBR components using a distributed computing environment
Authors: RAJAGOPAL A. , RAVICHANDRAN M. , MURALIDHER N. G. , SIVAKUMAR S. M.
Published: September 2003

In this work, the structural integrity of the critical components of the fast breeder reactor (FBR) that are subjected thermal striping is assessed using a fracture mechanics approach based on linear elastic fracture mechanics (LEFM). The structural integrity is assessed in terms of the actual life of the component for a particular difference between the hot and cold liquid temperatures at the critical mixing velocities. A generalized procedure is attempted for the computation of fatigue life. It is demonstrated in this work that the analysis procedure adopted is computationally very efficient. Green's function method is used for transient mode I crack propagation analysis. An inherent parallelism in the method is exploited for computational efficiency. A distributed computing environment is, therefore, used to demonstrate the effectiveness of Green's function method for crack propagation analysis for the kind of problem solved in this work. A simple idealization in the form of flat plate geometry is used in a numerical example to show the computational efficiency. The method shows a good scale‐up justifying the benefit of using a distributed computing environment given a large amount of input data for the thermal striping problem.
Relation between stimulus and response in frog olfactory receptor neurons in vivo
Authors: Rospars Jean‐Pierre , Lánský Petr , Duchamp André , Duchamp‐Viret Patricia
Published: September 2003

The spiking activity of receptor neurons was recorded extracellularly in the frog olfactory epithelium in response to four odourants applied at precisely controlled concentrations. A set of criteria was formulated to define the spikes in the response. Four variables – latency, duration, number of interspike intervals and frequency – were determined to quantify the responses. They were studied at the single neuron, neuron population and ciliary membrane levels. The dose–response curves were determined using specific functions and their characteristics were evaluated. The characteristic molar concentrations at threshold or at maximum duration and the characteristics of variables, e.g. minimum latency or maximum frequency, have asymmetric histograms with peaks close to the origin and long tails. Dynamic ranges have even more asymmetric histograms, so that a significant fraction of neurons presents a much wider range than their one‐decade peak. From these histograms, response properties of the whole neuron population can be inferred. In general, location along the concentration axis (thresholds), width (dynamic ranges) and heights of dose–response curves are independent, which explains the diversity of curves, prevents their global categorization and supports the qualitative coding of odourants. No evidence for odourant‐independent types of neurons was found. Finally, receptor activation and ciliary membrane conductance were reconstructed in the framework of a model based on firing data, known mucus biochemical and neuron morpho‐electrical characteristics. It is in agreement with independent determinations of Kd of odourant–receptor interaction and of conductance characteristics, and describes their statistical distributions in the neuron population.
Timecourse of striatal re‐innervation following lesions of dopaminergic SNpc neurons of the rat
Authors: Stanic D. , Finkelstein D. I. , Bourke D. W. , Drago J. , Horne M. K.
Published: September 2003

Previously we described the extent of sprouting that axons of the rat substantia nigra pars compacta (SNpc) undergo to grow new synapses and re‐innervate the dorsal striatum 16 weeks after partial lesions. Here we provide insights into the timing of events related to the re‐innervation of the dorsal striatum by regenerating dopaminergic nigrostriatal axons over a 104‐week period after partial SNpc lesioning. Density of dopamine transporter and tyrosine hydroxylase immunoreactive axonal varicosities (terminals) decreased up to 80% 4 weeks after lesioning but returned to normal by 16 weeks, unless SNpc lesions were greater than 75%. Neuronal tracer injections into the SNpc revealed a 119% increase in axon fibres (4 mm rostral to the SNpc) along the medial forebrain bundle 4 weeks after lesioning. SNpc cells underwent phenotypic changes. Four weeks after lesioning the proportion of SNpc neurons that expressed tyrosine hydroxylase fell from 90% to 38% but returned to 78% by 32 weeks. We discuss these phenotype changes in the context of neurogenesis. Significant reductions in dopamine levels in rats with medium (30–75%) lesions returned to normal by 16 weeks whereas recovery was not observed if lesions were larger than 75%. Finally, rotational behaviour of animals in response to amphetamine was examined. The clear rightward turning bias observed after 2 weeks recovered by 16 weeks in animals with medium (30–75%) lesions but was still present when lesions were larger. These studies provide insights into the processes that regulate sprouting responses in the central nervous system following injury.
The immunosuppressant mycophenolate mofetil attenuates neuronal damage after excitotoxic injury in hippocampal slice cultures
Authors: Dehghani Faramarz , Hischebeth Gunnar T. R. , Wirjatijasa Florentina , Kohl Angelika , Korf Horst‐Werner , Hailer Nils P.
Published: September 2003

In this study we investigated whether treatment with the immunosuppressant mycophenolate mofetil (MMF) has beneficial effects on neuronal damage after excitotoxic injury. Organotypic hippocampal slice culture (OHSC), lesioned by the application of N‐methyl‐d‐aspartate (NMDA) after 6 days in vitro, showed an improved preservation of the hippocampal cytoarchitecture after continuous treatment with MMF for 3 further days (10 or 100 µg/mL). Treatment with NMDA and MMF (100 µg/mL) reduced the number of damaged propidium iodide (PI)+ neurons by 50.7% and the number of microglial cells by 52%. Continuous treatment of lesioned OHSCs with MMF for 3 days almost abrogated the glial proliferative response, reflected by the 91.5% reduction in the number of bromo‐desoxy‐uridine (BrdU)‐labelled microglial cells and astrocytes. Microglial cells in MMF‐treated OHSCs contained fragmented nuclei, indicating apoptotic cell death, an effect which was also found in isolated microglial cells treated with MMF. The beneficial effect of MMF on neuronal survival apparently does not reflect a direct antiexcitotoxic effect, as short‐term treatment of OHSCs with NMDA and MMF for 4 h did not reduce the number of PI+ neurons. In conclusion, MMF inhibits proliferation and activation of microglia and astrocytes and protects neurons after excitotoxic injury.
The in vivo positional identity gene expression code is not preserved in neural stem cells grown in culture
Authors: Santa‐Olalla Jesús , Baizabal José‐Manuel , Fregoso Mariana , Del Carmen Cárdenas María , Covarrubias Luis
Published: September 2003

Neural stem cell specification depends on antero‐posterior (AP) and dorso‐ventral (DV) information provided during development. In the present study we identified similar neural stem cell (NSC) populations along the AP axis of the mouse central nervous system: the ‘early’ NSCs responsive to fibroblast growth factor‐2 and the ‘late’ NSCs responsive to epidermal growth factor (EGF). Gene expression analysis shows that AP and DV transcription factor code is not preserved in NSCs in culture. Neurospheres generated with EGF from different regions showed Emx2, En2 and Krox20 expression beyond their corresponding AP restricted areas (telencephalon, mesencephalon and rhomboencephalon, respectively). Hox genes were rarely expressed. DV markers such as Pax7 and Dbx1 were not expressed in neurosphere cells, whereas Pax6 and Nkx2.1 were highly expressed independently of the NSC source region. In general, this pattern was found under different culture conditions. We propose that signals surrounding NSCs determine their positional identity gene expression code, which may be relevant to establish their definitive fate.
Increase in Bcl‐2 phosphorylation and reduced levels of BH3‐only Bcl‐2 family proteins in kainic acid‐mediated neuronal death in the rat brain
Authors: Korhonen Laura , Belluardo Natale , Mudo Giuseppa , Lindholm Dan
Published: September 2003

Kainic acid induces excitotoxicity and nerve cell degeneration in vulnerable regions of rat brain, most markedly in hippocampus and amygdala. Part of the cell death following kainic acid is apoptotic as shown by caspase 3 activation and chromatin condensation. Here we have studied the regulation of pro‐ and anti‐apoptotic proteins belonging to the Bcl‐2 family in rat hippocampus and amygdala by kainic acid in relationship to ensuing neuronal death. The pro‐apoptotic protein Bax was up‐regulated in hippocampus 6 h after kainic acid administration. The increase in Bax was followed by the appearance of TdT‐mediated dUTP nick end labelling‐positive cells which were prominent at 24 h. Immunohistochemistry for active Bax revealed a punctated labelling of neurons in the CA3 and hilar regions of hippocampus as well as in amygdala. Double staining for NeuN, a marker for nerve cells, and TdT‐mediated dUTP nick end labelling showed that mainly neurons undergo degeneration after kainic acid treatment. In contrast to Bax, the pro‐apoptotic BH3‐only Bcl‐2 proteins Bim and Harakiri/DP5 were down‐regulated by kainic acid. This was also observed for the anti‐apoptotic proteins Bcl‐x and Bcl‐w. Immunoreactive Bcl‐2 was up‐regulated in hippocampus after kainic acid together with an increase in the phosphorylation of serine‐87 in Bcl‐2, suggesting a post‐transcriptional modification of the protein. This was confirmed using immunoprecipitation of total Bcl‐2 from hippocampus and amygdala which revealed an increase in serine‐87 phospho‐Bcl‐2 after kainic acid. Inhibition of the c‐jun N‐terminal protein kinase pathway reduced both serine‐87 phosphorylation and cell death after kainic acid. This indicates an important role of Bcl‐2 phosphorylation in controlling neuronal death after kainic acid. In contrast to the situation in trophic factor‐deprived neurons, no up‐regulation of Bim or Harakiri/DP5 proteins occurred after kainic acid, suggesting alternative pathways for regulation of cell death in excitotoxicity. The results indicate that not only the relative levels of Bcl‐2 family proteins but also conformation changes and post‐translational modifications contribute to neuronal death following kainic acid.
In vivo modulation of 5‐hydroxytryptamine release in mouse prefrontal cortex by local 5‐HT2A receptors: effect of antipsychotic drugs
Authors: Bortolozzi Analía , Amargós‐Bosch Mercè , Adell Albert , Díaz‐Mataix Llorenç , Serrats Jordi , Pons Sebastià , Artigas Francesc
Published: September 2003

In the rat, postsynaptic 5‐hydroxytryptamine2A receptors medial prefrontal cortex control the activity of the serotonergic system through changes in the activity of pyramidal neurons projecting to the dorsal raphe nucleus. Here we extend these observations to mouse brain. The prefrontal cortex expresses abundant 5‐ hydroxytryptamine2A receptors, as assessed by immunohistochemistry, Western blots and in situ hybridization procedures. The application of the 5‐hydroxytryptamine2A/2C agonist DOI (100 µm) by reverse dialysis in the medial prefrontal cortex doubled the local release of 5‐hydroxytryptamine. This effect was reversed by coperfusion of tetrodotoxin, and by the selective 5‐hydroxytryptamine2A receptor antagonist M100907, but not by the 5‐hydroxytryptamine2C antagonist SB‐242084. The effect of DOI was also reversed by prazosin (α1‐adrenoceptor antagonist), BAY × 3702 (5‐hydroxytryptamine1A receptor agonist), NBQX (α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole‐4‐propionate/kainic acid antagonist) and 1S,3S‐ACPD (mGluR II/III agonist), but not by dizocilpine (N‐methyl‐d‐aspartate antagonist). α‐Amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole‐4‐propionate mimicked the 5‐hydroxytryptamine elevation produced by DOI, an effect also reversed by BAY × 3702. Likewise, the coperfusion of classical (chlorpromazine, haloperidol) and atypical antipsychotic drugs (clozapine, olanzapine) fully reversed the 5‐hydroxytryptamine elevation induced by DOI. These observations suggest that DOI increases 5‐hydroxytryptamine release in the mouse medial prefrontal cortex through the activation of local 5‐hydroxytryptamine2A receptors by an impulse‐dependent mechanism that involves/requires the activation of local α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole‐4‐propionate receptors. This effect is reversed by ligands of receptors present in the medial prefrontal cortex, possibly in pyramidal neurons, which are involved in the action of antipsychotic drugs. In particular, the reversal by classical antipsychotics may involve blockade of α1‐adrenoceptors, whereas that of atypical antipsychotics may involve 5‐hydroxytryptamine2A receptors and α1‐adrenoceptors.
Different phosphatase‐dependent mechanisms mediate long‐term depression and depotentiation of long‐term potentiation in mouse hippocampal CA1 area
Authors: Jouvenceau Anne , Billard Jean‐Marie , Haditsch Ursula , Mansuy Isabelle M. , Dutar Patrick
Published: September 2003

Two types of synaptic depression have been described in the hippocampus, long‐term depression and depotentiation of long‐term potentiation known to recruit the serine/threonine protein phosphatases PP1, PP2A and PP2B (calcineurin). The contribution of each of these protein phosphatases is controversial. To examine the role of the Ca2+/calmodulin‐dependent protein phosphatase calcineurin in long‐term depression and depotentiation, we analysed the effect of genetically inhibiting calcineurin reversibly in the hippocampus, using the doxycycline‐dependent rtTA system in transgenic mice. We show that reducing calcineurin activity has no effect on long‐term depression but reversibly affects depotentiation. Consistently, the calcineurin inhibitor FK‐506 reproduces the depotentiation impairment observed in the mutant mice but does not affect long‐term depression in control animals. In contrast, the PP1/PP2A inhibitor okadaic acid fully blocks both long‐term depression and depotentiation. These data demonstrate that the nature of signalling cascades induced by synaptic activity depends on the initial synaptic state. While depression of potentiated synaptic responses requires activation of PP1/PP2A and/or calcineurin, depression of basal synaptic responses depends only on PP1/PP2A activation.
Kinase‐dependent loss of Na+ channel slow‐inactivation in rat CA1 hippocampal pyramidal cell dendrites after brief exposure to convulsants
Authors: Loftis Jordan L. , King Diana D. , Colbert Costa M.
Published: September 2003

Na+ channels in the dendrites of rat CA1 pyramidal neurons display a profound activity‐dependent inactivation, termed slow inactivation, that limits excitability in the dendrites even at low physiological rates of firing. The magnitude of this slow inactivation is powerfully modulated by a protein kinase C‐dependent process. Because activation of kinases is a rapid and common feature of a number of seizure models, we hypothesized that a loss of slow inactivation of Na+ channels might exacerbate other changes in excitability. Thus, we observed the effects of a brief (5 min) chemical convulsant treatment on Na+ currents and action potentials in hippocampal slices. We found that slow inactivation decreased significantly and remained decreased for at least 30 min after return to control conditions. Pretreatment with either chelerythrine, a protein kinase C inhibitor, or U0126, a mitogen‐activated protein kinase/extracellular signal regulated kinase kinase (MEK) inhibitor, blocked this reduction of slow inactivation. These results demonstrate that a brief period of hyperexcitability leads to a rapid, protein kinase‐dependent loss of slow inactivation of Na+ channels that would contribute to and perhaps prolong the hyperexcitable state.
Lesions of mediodorsal thalamus and anterior thalamic nuclei produce dissociable effects on instrumental conditioning in rats
Authors: Corbit Laura H. , Muir Janice L. , Balleine Bernard W.
Published: September 2003

Two experiments examined the effects of bilateral excitotoxic lesions of either the mediodorsal (MD) or anterior (ANT) thalamic nuclei on instrumental acquisition and performance, sensitivity to changes in the value of the instrumental outcome, and sensitivity to changes in the instrumental contingency. Rats were food deprived and trained to press two levers, each earning a unique food outcome (pellets or sucrose). All rats acquired the instrumental response although ANT lesions appear slightly to increase and MD lesions slightly to suppress instrumental performance. After training, specific satiety‐induced devaluation of one of the two instrumental outcomes produced a selective reduction in responding on the lever that in training had earned the now devalued outcome but only in the SHAM and ANT groups. In contrast, MD animals failed to show evidence of a selective devaluation effect when tested in extinction. Additionally, SHAM and ANT animals selectively decreased responding when one action‐outcome contingency was degraded, whereas MD animals reduced responding nonselectively on the two levers. Subsequent tests established that an inability to discriminate between either the two actions or the two outcomes cannot account for the lack of selective responding observed in the MD animals. Together these data suggest that MD lesions produce a profound deficit in the ability of rats to utilize specific action–outcome associations and appear to render rats relatively insensitive to the causal consequences of their instrumental actions. In contrast, far from producing a deficit, ANT lesioned rats were as sensitive to the effects of these behavioural manipulations as the sham lesioned controls.
Activation of retinoic acid signalling after sciatic nerve injury: up‐regulation of cellular retinoid binding proteins
Authors: Zhelyaznik Nina , Schrage Kirsten , McCaffery Peter , Mey Jörg
Published: September 2003

In mammalian peripheral nerves a crush lesion causes interactions between injured neurons, Schwann cells and haematogenous macrophages that can lead to successful axonal regeneration. We suggest that the transcriptional activator retinoic acid (RA), takes part in gene regulation after peripheral nerve injury and that RA signalling is activated via the cellular retinoic acid binding protein (CRABP)‐II and cellular retinol binding protein (CRBP)‐I. With RT‐PCR and immunoblotting all necessary components of the RA signalling pathway were detected in the sciatic nerve of adult rats. These are retinoic acid receptors, retinoid X receptors, the retinoic acid synthesizing enzymes RALDH‐1, RALDH‐2, and RALDH‐3, in addition, the cellular retinoid binding proteins CRBP‐I, CRABP‐I and CRABP‐II. Enzyme activity of RALDH‐2 was detectable in the nerve, and using a transgenic reporter mouse we found local activation of RA responsive elements in the regenerating nerve. Sciatic nerve crush as well as transection resulted in a more than 10‐fold up‐regulation of CRBP‐I, which is thought to facilitate the synthesis of RA. Both kinds of injury also caused a 15‐fold increase in transcript and protein concentration of CRABP‐II, a possible mediator of RA transfer to its nuclear receptors.
Distinct receptors and different transduction mechanisms for ATP and adenosine at the frog motor nerve endings
Authors: Sokolova E. , Grishin S. , Shakirzyanova A. , Talantova M. , Giniatullin R.
Published: September 2003

Corelease of ATP with ACh from motor endings suggests a physiological role for ATP in synaptic transmission. We previously showed that, on skeletal muscle, ATP directly inhibited ACh release via presynaptic P2 receptors. The receptor identification (P2X or P2Y) and its transduction mechanism remained, however, unknown. In the present study using the voltage‐clamp technique we analyzed the properties of presynaptic ATP receptors and subsequent effector mechanisms. ATP or adenosine presynaptically depressed multiquantal end‐plate currents, with longer latency for ATP action. ATPγS, agonist at P2X receptors, or Bz‐ATP, agonist at P2X7 receptors, were ineffective. The action of ATP was prevented by suramin and unchanged by PPADS or TNP‐ATP, antagonists of P2X receptors, or RB‐2, a blocker of certain P2Y receptors. The depressant action of ATP was reproduced by UTP, metabotropic P2Y receptor agonist. Pertussis toxin (PTX), antagonist of Gi/o‐proteins, and inhibitors of phosphatidylcholine specific PLC (D609) and PKC (staurosporine or chelerythrine) prevented the effect of ATP while blockers of PLA2 (OBAA) and COX (aspirin or indomethacin) attenuated it. Inhibitors of phosphatidylinositide‐specific PLC (U73122), guanylylcyclase (ODQ), PKA (Rp‐cAMPS) or PLD (1‐butanol) did not affect the action of ATP. No inhibitor of second messengers (except PTX) changed the action of adenosine. Our data indicate, for motor nerve endings, the existence of inhibitory P2Y receptors coupled to multiple intracellular cascades including phosphatidylinositide‐specific PLC/PKC/PLA2/COX. This divergent presynaptic P2 signalling (unlike the single effector mechanism for P1 receptors) could provide feedback inhibition of transmitter release and perhaps be involved in presynaptic plasticity.
Interleukin‐1β enhances non‐rapid eye movement sleep when microinjected into the dorsal raphe nucleus and inhibits serotonergic neuronsin vitro
Authors: Manfridi Alfredo , Brambilla Dario , Bianchi Susanna , Mariotti Maurizio , Opp Mark R. , Imeri Luca
Published: September 2003

Interleukin‐1 (IL‐1) and IL‐1 receptors are constitutively expressed in normal brain. IL‐1 increases non‐rapid eye movements (NREM) sleep in several animal species, an effect mediated in part by interactions with the serotonergic system. The site(s) in brain at which interactions between IL‐1 and the serotonergic system increase NREM sleep remain to be identified. The dorsal raphe (DRN) is the origin of the major ascending serotonergic pathways to the forebrain, and it contains IL‐1 receptors. This study examined the hypothesis that IL‐1 increases NREM sleep by acting at the level of the DRN. IL‐1β (0.25 and 0.5 ng) was microinjected into the DRN of freely behaving rats and subsequent effects on sleep–wake activity were determined. IL‐1β 0.5 ng increased NREM sleep during the first 2 h post‐injection from 33.5 ± 3.7% after vehicle microinjection to 42.9 ± 3.0% of recording time. To determine the effects of IL‐1β on electrophysiological properties of DRN serotonergic neurons, intracellular recordings were performed in a guinea‐pig brain stem slice preparation. In 26 of 32 physiologically and pharmacologically identified serotonergic neurons, IL‐1β superfusion (25 ng/mL) decreased spontaneous firing rates by 50%, from 1.6 ± 0.2 Hz (before IL‐1β superfusion) to 0.8 ± 0.2 Hz. This effect was reversible upon washout. These results show that IL‐1β increases NREM sleep when administered directly into the DRN. Serotonin enhances wakefulness and these novel data also suggest that IL‐1β‐induced enhancement of NREM sleep could be due in part to the inhibition of DRN serotonergic neurons.
c‐Fos and peptide immunoreactivities in the central extended amygdala of morphine‐dependent rats after naloxone‐precipitated withdrawal
Authors: Veinante Pierre , Stoeckel Marie‐Elisabeth , Lasbennes François , Freund‐Mercier Marie‐José
Published: September 2003

The central extended amygdala, a forebrain macrostructure, may represent a common substrate for acute drug reward and the dysphoric effects of drug withdrawal. To test its involvement during opiate withdrawal, we studied the distribution of c‐Fos immunoreactive neurons, in relation to their neuropeptide content, in brain sections from morphine‐dependent or naive rats, killed 90 min after naloxone or saline intraperitoneal injection. Naloxone treatment in naive rats induced a slight increase in c‐Fos immunoreactivity in the central amygdaloid nucleus, the lateral bed nucleus of the stria terminalis and the interstitial nucleus of the posterior limb of the anterior commissure. In morphine‐dependent rats, naloxone injection significantly increased the number of c‐Fos‐positive neurons in these structures as well as in the majority of the other central extended amygdala components. Double immunocytochemistry was used to determine the neurochemical nature of c‐Fos‐positive neurons in the central extended amygdala. Corticotropin‐releasing factor‐ and methionine‐enkephakin‐immunoreactive neurons displayed c‐Fos immunoreactivity in naive rats after naloxone injection, whereas only enkephalinergic neurons were found to be c‐Fos positive in morphine‐dependent rats after naloxone injection. The possible involvement of the corticotropin‐releasing factor system during withdrawal is discussed. These results suggest that the whole central extended amygdala is activated during opiate withdrawal, with a lateral to medial decreasing gradient, and emphasize the role of peptidergic systems in this morphofunctional continuum.
Tissue inhibitor of metalloproteinases‐3 and matrix metalloproteinase‐3 regulate neuronal sensitivity to doxorubicin‐induced apoptosis
Authors: Wetzel M. , Rosenberg G. A. , Cunningham L. A.
Published: September 2003

Metalloproteinase activity at the cell surface influences cellular sensitivity to extrinsic death vs. survival signals in a variety of cell types, through proteolytic shedding of cell surface signalling molecules. Tissue inhibitor of metalloproteinases‐3 (TIMP‐3) is a unique natural metalloproteinase inhibitor that plays a pro‐apoptotic role through its ability to inhibit metalloproteinases that proteolytically cleave death receptors and their ligands from the cell surface. To study the convergence of metalloproteinase activity and death receptor signalling in neurons, we established an in vitro model of neuronal apoptosis utilizing the chemotherapeutic drug, doxorubicin (Dox). Primary cultures established from embryonic rat cerebral cortices displayed robust and selective neuronal apoptosis in response to Dox, an effect that was dependent on the activation of the death receptor, Fas. We demonstrate that both TIMP‐3 and matrix metalloproteinase‐3 (MMP‐3) are constitutively expressed by primary cortical neurons in culture, and selectively modulated Fas‐mediated neuronal apoptosis induced by Dox. Metalloproteinase inhibition by TIMP‐3 was found to be necessary for Dox‐induced neuronal death, whereas addition of active MMP‐3 markedly attenuated apoptosis and diminished Fas–Fas ligand interaction at the cell surface. These observations implicate a physiological role for the balance of TIMP‐3 and MMP‐3 activity at the neuronal surface in regulating death receptor sensitivity. The convergence of metalloproteinase activity and death receptor signalling at the cell surface may influence neuronal cell death vs. survival decisions.
Compensatory dendritic growth of CA1 pyramidal cells following growth impairment in the neonatal period
Authors: Groc Laurent , Petanjek Zdravko , Gustafsson Bengt , Ben‐Ari Yehezkel , Khazipov Roustem , Hanse Eric
Published: September 2003

In the neonatal rat, inhibition of hippocampal neural activity in vivo by tetanus toxin results in a severe growth impairment of the basal dendrites of CA1 pyramidal cells. Here we tested whether this early growth impairment results in a permanent reduction of the basal dendritic tree or whether recovery processes are recruited later in development when synaptic activity has fully recovered. Quantitative analysis of dendritic parameters and spine density from reconstructed CA1 pyramidal cells showed that young adult CA1 pyramidal (postnatal day 31–34) cells that were exposed to activity deprivation in the neonatal period were almost indistinguishable from control cells of the same age. These results suggest that the early hippocampal activity controls the growth rate but is not necessary for the generation of an adult normal basal dendritic tree.
Plasma membrane and vesicular glutamate transporter mRNAs/proteins in hypothalamic neurons that regulate body weight
Authors: Collin Maria , Bäckberg Matilda , Ovesjö Marie‐Louise , Fisone Gilberto , Edwards Robert H. , Fujiyama Fumino , Meister Björn
Published: September 2003

After synaptic release, glutamate is taken up by the nerve terminal via a plasma membrane‐associated protein termed excitatory amino acid transporter 3 (EAAT3). Following entry into the nerve terminal, glutamate is pumped into synaptic vesicles by a vesicular transport system. Three different vesicular glutamate transporter proteins (VGLUT1‐3) representing unique markers for glutamatergic neurons were recently characterized. The presence of EAAT3, glutaminase and VGLUT1‐3 was examined in mouse, rat and rabbit species at mRNA and protein levels in hypothalamic neurons which are involved in the regulation of body weight using in situ hybridization and immunohistochemistry. EAAT3 and glutaminase mRNAs were demonstrated in all parts of the arcuate nucleus in the dorsomedial and ventromedial hypothalamic nuclei and lateral hypothalamic area. VGLUT1 mRNA was present in the magnocellular lateral hypothalamic nucleus. VGLUT2 mRNA was demonstrated in a subpopulation of neurons in the arcuate nucleus and in the ventromedial and dorsomedial hypothalamic nuclei and lateral hypothalamic area. Few VGLUT3 mRNA expressing neurons were scattered throughout the medial and lateral hypothalamus. EAAT3‐like immunoreactivity (‐li) was demonstrated in glutamate, neuropeptide Y (NPY), agouti‐related peptide (AGRP), pro‐opiomelanocortin (POMC), cocaine and amphetamine‐regulated transcript (CART), melanin‐concentrating hormone and orexin‐immunoreactive (‐ir) neurons. VGLUT2‐li could only be demonstrated in POMC‐ and CART‐ir neurons of the ventrolateral arcuate nucleus. The results show that key neurons involved in regulation of energy balance are glutamatergic and/or densely innervated by glutamatergic nerve terminals. Whereas orexigenic NYP/AGRP neurons situated in the ventromedial part of the arcuate nucleus are mainly GABAergic, it is shown that several anorexigenic POMC/CART neurons of the ventromedial arcuate nucleus are most likely glutamatergic.
Gene expression of AMPA‐type glutamate receptor subunits in rod‐type ON bipolar cells of rat retina
Authors: Kamphuis Willem , Dijk Frederike , O'Brien Brendan J.
Published: September 2003

The retinal rod bipolar cell type is involved in the sign‐inverting depolarizing ON‐type response to light. This response is mediated by the metabotropic glutamate receptor type 6 (mGluR6) expressed on the rod bipolar dendrites. In a previous immunocytochemical study, an unexpected colocalization was reported [W. Kamphuis et al. (2003) J. Comp. Neurol., 455, 172–186] of mGluR6 with the ionotropic AMPA‐type glutamate receptor subunit GluR2 in rod bipolar cells of rat retina. The aim of the present study was to investigate whether expression of both genes could be found at the single‐cell level. Two approaches were followed. (i) Retinal cells were isolated by enzymatic and mechanical treatment. Single cells with a bipolar morphology were harvested, subjected to multiplex PCR with protein kinase C (PKC)‐, mGluR6‐ and GluR1–4‐specific primers, followed by a real‐time quantitative PCR assay. Of 23 studied cells, 74% expressed PKC and 87% expressed mGluR6. Using the presence of both transcripts as the criterion for a rod bipolar cell signature (n = 15), 73% of these cells expressed GluR2, with a minor contribution of GluR1 (20%), GluR3 (7%), and GluR4 (20%). Quantification of the transcript levels demonstrated that mGluR6 and GluR2 genes are expressed at similar levels in rod ON‐type bipolar cells. (ii) Rod bipolar cells were identified in retinal sections by immunolabelling with a protein kinase C antibody and isolated using laser pressure catapulting (LPC). Quantitative PCR was employed to assess gene expression levels of reference genes, PKCα, mGluR6 and the GluR subunits. However, in samples from PKCα‐immunopositive somata no significant enrichment of PKCα transcript levels was observed when compared with control samples from immunonegative somata. We conclude that this approach lacks sufficient spatial specificity. In conclusion, the results show coexpression of mGluR6 and GluR2 in rod bipolar cells; this is in good agreement with the results of previous immunocytochemical studies. The functional implications of AMPA‐type glutamate receptors for ON‐type rod bipolar‐mediated signal transduction remains to be elucidated.
Morphometrical analysis of dendritic arborization in axotomized retinal ganglion cells
Authors: Germain Francisco , Fernández Eduardo , De La Villa Pedro
Published: September 2003

It has been reported that section of the optic nerve in mammals causes death in >90% of the retinal ganglion cells (RGCs). The cells which survive the section experience an irreparable loss of many of their dendritic segments and a rapid retraction of the dendritic tree. However, some growth cones and abnormal processes have been also reported. Our aim was to make a quantitative study of the morphological changes found in rabbit RGCs after optic nerve section. The morphometrical analysis of the RGCs which survived the axotomy showed an increase in the diameter of the soma and a significant increase in the area of the dendritic field; also, the length of the dendritic segments was significantly longer in axotomized RGCs than in control cells. Terminal dendritic segments (T) and preterminal segments (PT) were both measured in control and axotomized cells; the length ratio of T : PT segments was significantly greater in the axotomized cells than in the controls. We conclude that RGCs which survived the axotomy experienced a significant growth of their terminal dendritic branches.
Bicarbonate‐dependent depolarizing potentials in pyramidal cells and interneurons during epileptiform activity
Authors: Perez Velazquez Jose L.
Published: September 2003

Experimental and theoretical evidence indicates that GABAergic neurotransmission is fundamental for the synchronization of neuronal activity. In particular, the role of GABA in epileptiform activity has received increased attention due to, among others, the fact that the GABA‐mediated potentials can be depolarizing, and hence excitatory, in some circumstances. Evidence is presented here that bicarbonate efflux via GABAA receptors in interneurons and pyramidal cells of the CA1 hippocampal area contribute to depolarizing GABAA‐mediated potentials in an in vitro nonpharmacological seizure‐like model of status epilepticus. Seizure‐like and interictal activity was evoked in rat horizontal hippocampal slices using a superfusing solution with low magnesium concentration (0.5–0.9 mm). Whole‐cell recordings from stratum oriens–alveus interneurons and CA1 pyramidal cells revealed that, during epileptiform activity, some GABAA‐mediated potentials were depolarizing, and were suppressed by the carbonic anhydrase inhibitor ethoxyzolamide as well as by the GABAA‐receptor antagonist bicuculline. These observations indicate that the depolarizing potentials observed during epileptiform activity reflect both glutamatergic and GABAA‐receptor‐mediated activity, and adds further support for the important role of GABAergic interneurons in promoting long‐range synchronization.
Co‐localization of brain‐derived neurotrophic factor (BDNF) and wild‐type huntingtin in normal and quinolinic acid‐lesioned rat brain
Authors: Fusco Francesca R. , Zuccato Chiara , Tartari Marzia , Martorana Alessandro , De March Zena , Giampà Carmela , Cattaneo Elena , Bernardi Giorgio
Published: September 2003

Loss of huntingtin‐mediated brain‐derived neurotrophic factor (BDNF) gene transcription has been described in Huntington's disease (HD) [Zuccato et al. (2001) Science, 293, 493–498]. It has been shown that BDNF is synthesized in the pyramidal layer of cerebral cortex and released in the striatum [Altar et al. (1997) Nature, 389, 856–860; Conner et al. (1997) J. Neurosci., 17, 2295–2313]. Here we show the cellular localization of BDNF in huntingtin‐containing neurons in normal rat brain; our double‐label immunofluorescence study shows that huntingtin and BDNF are co‐contained in ≈99% of pyramidal neurons of motor cortex. In the striatum, huntingtin is expressed in 75% of neurons containing BDNF. In normal striatum we also show that BDNF is contained in cholinergic and in NOS‐containing interneurons, which are relatively resistant to HD degeneration. Furthermore, we show a reduction in huntingtin and in BDNF immunoreactivity in cortical neurons after striatal excitotoxic lesion. Our data are confirmed by an ELISA study of BDNF and by a Western blot analysis of huntingtin in cortex of quinolic acid (QUIN)‐lesioned hemispheres. In the lesioned striatum we describe that the striatal subpopulation of cholinergic neurons, surviving degeneration, contain BDNF. The finding that BDNF is contained in nearly all neurons that contain huntingtin in the normal cortex, along with the reduced expression of BDNF after QUIN injection of the striatum, shows that huntingtin may be required for BDNF production in cortex.
Acute stress increases calcium current amplitude in rat hippocampus: temporal changes in physiology and gene expression
Authors: Joëls M. , Velzing E. , Nair S. , Verkuyl J. M. , Karst H.
Published: September 2003

Activation of hippocampal glucocorticoid receptors in vitro increases calcium current amplitude through a process requiring DNA binding of receptor homodimers. We here investigated (i) whether similar increased calcium currents also occur following in vivo glucocorticoid receptor activation due to stress and (ii) if so, whether this can be explained by increased expression of calcium channel subunits. Rats were exposed to a novelty stress; some of the animals were pretreated with a glucocorticoid receptor antagonist. In subsequently prepared hippocampal slices, calcium currents were recorded from identified CA1 pyramidal neurons, after which RNA was collected, linearly amplified and hybridized with cDNA clones. Glucocorticoid receptor activation due to novelty exposure was associated with large total peak calcium currents and high‐threshold noninactivating currents. Low‐threshold calcium currents were not affected. Large total peak and noninactivating current amplitudes were also seen when animals received a more severe stressor, i.e. additional ether exposure. In the stressed groups, the total peak and high‐threshold calcium current gradually increased with time resulting in a significant enhancement at ≥3 h after stress exposure. In the same cells, the summated (relative) RNA expression of various α1 calcium channel subunits was only transiently enhanced, prior to the functional changes. These data indicate that in vivo activation of glucocorticoid receptors due to stress gradually increases specific calcium current components. Prior to the functional change, increased expression of calcium channel subunits was observed, suggesting that the enhanced function could be explained by transcriptional regulation of the channels.
Potent mammalian cerebroprotection and neuronal cell death inhibition are afforded by a synthetic antioxidant analogue of marine invertebrate cell protectant ovothiols
Authors: Vamecq Joseph , Maurois Pierre , Bac Pierre , Bailly Fabrice , Bernier Jean‐Luc , Stables James P. , Husson Isabelle , Gressens Pierre
Published: September 2003

Implicit strategies for neuroprotection in the adult brain include GABAAreceptor activation,N‐methyl‐d‐aspartate receptor and sodium voltage‐gated channel inhibition. Ironically, these same targets may be harmful to the immature or developing brain. Protection has been demonstrated for both immature and mature brain with the use of a synthetic ovothiol analogue. The following beneficial effects have been demonstrated in mice: protection against audiogenic seizures, brain structures with clear‐cut delineation of ibotenate‐challenged white and grey matter lesions along with exceptional early and delayed protections, and potent cerebral cell death inhibition. The compound lacks both GABAergic activity and sodium channel blocker properties, which may help explain the lack of toxicity normally expressed in an immature brain utilizing these agents [J.W. Olney (2002) Neurotoxicology, 93, 1–10]. The oxidized form of the compound is virtually devoid of antioxidant activity.In vivoit exhibits cerebroprotective properties similar to those of reduced compounds endowed with antioxidant properties. This unexpected finding has prompted an extensivein vitroexploration of underlying molecular mechanisms that have led to the identification of several recycling mechanisms consistent with non rate‐limiting conversion of oxidized to reduced compound forms. Taken as a whole, this work offers an unique combinedin vitroandin vivosupport that: (i) antioxidant therapy, here engineered from marine invertebrate egg protectants, may be a valuable strategy in protecting both mammalian adult and developing brain; and (ii) recycling (thiol‐disulphide exchange) properties of the oxidized form of an antioxidant compound are as important as the antioxidant potential exhibited by a bioactive reduced antioxidant in certain neuroprotective processes.
Nicotine recruits a local glutamatergic circuit to excite septohippocampal GABAergic neurons
Authors: Wu Min , Hajszan Tibor , Leranth Csaba , Alreja Meenakshi
Published: September 2003

Tonic impulse flow in the septohippocampal GABAergic pathway is essential for normal cognitive functioning and is sustained, in part, by acetylcholine (ACh) that is released locally via axon collaterals of septohippocampal cholinergic neurons. Septohippocampal cholinergic neurons degenerate in Alzheimer's disease and other neurodegenerative disorders. While the importance of the muscarinic effects of ACh on septohippocampal GABAergic neurons is well recognized, the nicotinic effects of ACh remain unstudied despite the reported benefits of nicotine on cognitive functioning. In the present study, using electrophysiological recordings in a rat brain slice preparation, rapid applications of nicotine excited 90% of retrogradely labelled septohippocampal GABA‐type neurons with an EC50 of 17 µm and increased the frequency of spontaneously occurring, impulse‐dependent fast GABAergic and glutamatergic synaptic currents via the α4β2‐nicotinic receptor. Interestingly, tetrodotoxin blocked all effects of nicotine on septohippocampal GABAergic type neurons, suggesting involvement of indirect mechanisms. We demonstrate that the effects of nicotine on septohippocampal GABA‐type neurons involve recruitment of a novel, local glutamatergic circuitry as (i) Group I metabotropic glutamatergic receptor antagonists reduced the effects of nicotine; (ii) the number of nicotine responsive neurons was significantly reduced in recordings from slices that had been trimmed so as to reduce the number of glutamate‐containing neurons within the slice preparation; (iii) in light and ultrastructural double immunocytochemical labelling studies vesicular glutamate 2 transporter immunoreactive terminals made synaptic contacts with parvalbumin‐immunoreactive septohippocampal GABAergic neurons. The discovery of a local glutamatergic circuit within the septum may provide another avenue for restoring septohippocampal GABAergic functions in neurodegenerative disorders associated with a loss of septohippocampal cholinergic neurons.