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W Kamphuis

Publications and source records attributed to W Kamphuis.

At least 19 recordsLinked to original sources
PubMed2002

Intravitreal injection of adeno-associated viral vectors results in the transduction of different types of retinal neurons in neonatal and adult rats: a comparison with lentiviral vectors.

Replication-deficient viral vectors encoding the marker gene green fluorescent protein (GFP) were injected into the vitreous of newborn, juvenile (P14), and adult rats. We tested two different types of modified virus: adeno-associated viral-2-GFP (AAV-GFP) and lentiviral-GFP vectors (LV-GFP). The extent of retinal cell transduction in different-aged animals was compared 7, 21, and 70 days after eye injections. At all postinjection times, LV-GFP transduction was mostly limited to pigment epithelium and cells in sclera and choroid. In contrast, transduction of large numbers of neural retinal cells was seen 21 and 70 days after AAV-GFP injections. AAV-GFP predominantly transduced neurons, although GFP-positive Müller cells were seen. All neuronal classes were labeled, but the extent of transduction for a given class varied depending on injection age. After P0 injections about 50% of transduced cells were photoreceptors and 30-40% were amacrine or bipolar cells. After adult injections 60-70% of transduced cells were retinal ganglion cells. In adults many GFP-positive retinal axons were traced through the optic nerve/tract and terminal arbors were visualized in central targets.

Adenoviridae↗
PubMed2001

Prostanoid receptor gene expression profile in human trabecular meshwork: a quantitative real-time PCR approach.

PURPOSE: To assess the expression pattern of prostanoid receptor-encoding genes in trabecular meshwork (TM) of human donor eyes. METHODS: Disposed human donor eyes (n = 10) were obtained from the Cornea Bank, Amsterdam. The TM was dissected from the scleral tissue and homogenized in lysis buffer, and total RNA was isolated. The RNA was converted into cDNA and used as a template for noncompetitive quantitative real-time polymerase chain reaction (PCR) using green fluorescent dye to quantify the accumulation of double-stranded PCR product. Specific primers for four housekeeping genes and DP, EP(1), EP(2), EP(3,) EP(4), FP, IP, and TP receptor-encoding transcripts were developed and tested for their efficiency. RESULTS: The characterized expression profile was highly reproducible in all samples, with the EP(2) receptor-encoding transcript in the highest abundance, followed by FP, TP, IP, and EP(4) at levels that were approximately 10 to 15 times lower than that of the EP(2) subtype. DP and EP(3) were at the lowest levels, which were, on average, 45 times and 228 times lower than EP(2), respectively. CONCLUSIONS: These data show that all prostanoid receptors are expressed at different levels in human TM tissue. Because the gene expression of the EP(2) receptor is, on average, 15 times more abundant than that of the EP(4) receptor, it may be expected that the increase in flow and cAMP levels in response to the activation of the EP receptors by application of prostaglandin E(1) (PGE(1)), is primarily mediated by the EP(2) receptor. These data should be considered when designing prostanoid receptor mimetics intended to enhance the aqueous humor outflow through the TM and Schlemm's canal.

Actins↗
PubMed2000

Immunocytochemical localization of the glutamate transporter GLT-1 in goldfish (Carassius auratus) retina.

Glutamate is the major excitatory neurotransmitter in the retina of vertebrates. Electrophysiological experiments in goldfish and salamander have shown that neuronal glutamate transporters play an important role in the clearance of glutamate from cone synaptic clefts. In this study, the localization of the glutamate transporter GLT-1 has been investigated immunocytochemically at the light and electron microscopical levels in the goldfish retina using a GLT-1-specific antibody. GLT immunoreactivity (IR) was observed at the light microscopical level in Müller cells, bipolar cells, the outer plexiform layer (OPL), and the inner plexiform layer (IPL). At the electron microscopical level, membrane-bound and cytoplasmic GLT-IR in the OPL was located in finger-like protrusions of the cone terminal located near the invaginating postsynaptic processes of bipolar and horizontal cells. GLT-IR was not observed in the vicinity of synaptic ribbons. This location of GLT-1 allows modulation of the glutamate concentration in the synaptic cleft, thereby shaping the dynamics of synaptic transmission between cones and second-order neurons. In the inner IPL, GLT-IR was observed in the cytoplasm and was membrane bound in mixed rod/cone bipolar cell terminals and cone bipolar cell terminals. The membrane-bound GLT-1 was generally observed at some distance from the synaptic ribbon. The morphology of the bipolar cell terminal together with the localization of GLT-1 suggests that at least these glutamate transporters are not primarily involved in rapid uptake of glutamate release by the bipolar cells. The GLT-IR in the cytoplasm of Müller cells was located throughout the entire goldfish retina from the outer limiting membrane to the inner limiting membrane. The location of GLT-1 in Müller cells is consistent with the role of Müller cells in converting glutamate to glutamine.

ATP-Binding Cassette Transporters↗
PubMed2000

Immunohistochemical localization of substance P and substance P receptor (NK1) in the olivary pretectal nucleus of the rat.

The olivary pretectal nucleus (OPN) is the first central nucleus in the pupillary light reflex arc (PLR). Substance P (SP) is a neuropeptide present in the OPN. The present immunohistochemical study, performed at the ultrastructural level, aimed to determine the synaptic localization of SP and SP receptor in the OPN. Three types of SP-positive terminals were found. The most abundant type was of retinal origin, characterized by electron-lucent mitochondria and round vesicles, organized in glomerular structures, making asymmetric synaptic contacts with dendrites, and profiles containing pleomorphic vesicles, also making synaptic contacts with dendrites. The second type of SP-immunoreactive terminal contained electron-dense mitochondria and pleomorphic vesicles. This type made symmetric synaptic contacts and may originate from the ventral part of the lateral geniculate nucleus. The third type of SP-immunoreactive terminals contained electron-dense mitochondria, clear round vesicles, and made an asymmetric synaptic contact. This type originates from the contralateral OPN. SP receptors of the NK1 subtype were revealed to be on dendrites and were part of the glomerular-like arrangement. On account of the present observations, it can be concluded that retinal projections to the OPN use SP as a neuromodulator and synapse on NK1 receptor-containing dendrites of large neurons projecting to the Edinger-Westphal nucleus. Since SP also modulates the parasympathetic component of the PLR, we postulate that SP plays a modulating role in all components of the PLR.

Animals↗
PubMed2000

Expression and localization of ionotropic glutamate receptor subunits in the goldfish retina--an in situ hybridization and immunocytochemical study.

The expression and distribution of AMPA, kainate and NMDA glutamate receptor subunits was studied in the goldfish retina. For the immunocytochemical localization of the AMPA receptor antisera against GluR2, GluR2/3 and GluR4 were used, and for in situ hybridization rat specific probes for GluR1 and GluR2 and goldfish specific probes for GluR3 and GluR4 were used. The localization of the low affinity kainate receptor and NMDA receptor was studied using antisera against GluR5-7 and NR1. All AMPA receptor subtypes were demonstrated to be present in the goldfish retina both by immunocytochemistry and in situ hybridization. In situ hybridization revealed expression of all AMPA receptors subunit at the inner border of the INL. Only GluR3 was also strongly expressed in the outer border of the INL. Some of the ganglion cells displayed a strong signal for GluR1, GluR3 and GluR4. GluR1-immunoreactivity was present in subsets of bipolar, amacrine, and ganglion cells. GluR2 and GluR2/3-immunoreactivity was mainly localized in the outer plexiform layer. GluR2 and GluR2/3-immunoreactivity are associated with the photoreceptor synaptic terminals. GluR4-immunoreactivity is present on Müller cells in the inner retina and on dendrites of bipolar cells in the OPL, whereas GluR5-7-immunoreactivity was prominently present on horizontal cell axon terminals. Finally, NR1-immunoreactivity was confined to amacrine cells, the inner plexiform layer and ganglion cells. This study shows that there is a strong heterogeneity of glutamate receptor subunit expression in the various layers of the retina. Of the AMPA receptor subunits GluR3 seems to be expressed the most widely in all layers with strong glutamatergic synaptic interactions whereas all the other subunits seem to have a more restricted expressed pattern.

Animals↗
PubMed1998

Expression patterns of voltage-dependent calcium channel alpha 1 subunits (alpha 1A-alpha 1E) mRNA in rat retina.

The transcript levels of the genes encoding for the different alpha1 (alpha1A-alpha1E) subunits of voltage-dependent calcium channels (VDCCs) were studied in the retina of the rat using RT-PCR, Northern blotting, and in situ hybridization. Abundant expression of alpha1A and alpha1B was found with RT-PCR and on Northern blots of total retina RNA, corresponding with high expression levels in all nuclear layers (outer and inner nuclear layers and the ganglion cell layer) of the retina. VDCC alpha1D mRNA was also present in all nuclear layers of the retina but at less abundant levels than alpha1A or alpha1B. Expression level of alpha1C in the retina was low as deduced from a faint Northern blot signal and a moderate yield after PCR amplification. VDCC alpha1E specific amplification of retinal cDNA yielded a longer product (designated alpha1E-L) than obtained from the hippocampus. Nucleotide sequencing of this PCR product revealed a 129 bp insert which is largely homologous (97%) with a previously described insert in the same position in human alpha1E cDNA. In situ hybridization in rat brain showed a differential expression pattern of the long and short variants of alpha1E mRNA. Northern blotting of retinal RNA confirmed the absence of the short variant (alpha1E-S), while alpha1E-L was present at low levels. In situ hybridization detected a significant level of expression of alpha1E-L in the inner nuclear layer. The prevalent expression of alpha1A and alpha1B, and to a lesser extent, of alpha1D, indicates that P/Q-, N-, and L-type calcium currents play a prominent role in the various cell types involved in the retinal signal-transduction pathway. The absence of alpha1C transcript in the retina suggests that the slowly inactivating L-type calcium currents involved in neurotransmitter release from the terminals of photoreceptors and bipolar cells may be encoded by the alpha1D isoform.

Alternative Splicing↗
PubMed1997

Pseudo-immunolabelling with the avidin-biotin-peroxidase complex (ABC) due to the presence of endogenous biotin in retinal Müller cells of goldfish and salamander.

Immunodetection techniques are dependent on enzyme-protein conjugates for the visualisation of antigen-antibody complexes. One of the most widely used is the avidin-biotin-peroxidase complex (ABC) method. The present study demonstrates that direct treatment of goldfish and salamander retinal sections with ABC, followed by an incubation with the chromogenic substrate 3,3-diaminobenzidine tetrahydrochloride (DAB) and H2O2, manifested a punctate staining pattern across the neural retinae, presumably through binding of avidin to endogenous biotin. Incubation with a primary antiserum against biotin followed by immunoprocessing with the peroxidase--anti-peroxidase (PAP) method showed a pattern similar to the punctuate framework as detected with solo ABC-treated sections. Moreover, the ABC-DAB/H2O2 mediated pattern corresponded to the spatial orientation of Müller cells as identified by GFAP immunostaining. These findings indicate the presence of endogenous biotin in Müller cells and calls for caution in the application of the ABC method in immunotechniques in retinal research.

Animals↗
PubMed1997

Changes in voltage-dependent calcium channel alpha1-subunit mRNA levels in the kindling model of epileptogenesis.

The establishment of a focus of epileptiform activity in the hippocampus of the rat, using the kindling paradigm, leads to enhanced voltage-dependent calcium conductance of CA1 pyramidal neurones (G.C. Faas, M. Vreugdenhil, W.J. Wadman, Calcium currents in pyramidal CA1 neurones in vitro after kindling epileptogenesis in the hippocampus of the rat, Neuroscience 75 (1996) 57-67; M. Vreugdenhil, W.J. Wadman, Kindling-induced long-lasting enhancement of calcium in hippocampal CA1 area of the rat: relation to calcium-dependent inactivation, Neuroscience 59 (1994) 105-114). Using semi-quantitative in situ hybridization techniques, we investigated whether these changes were associated with an altered expression of the genes that encode for the alpha1A-E-subunits of the voltage-dependent calcium channels (VDCC). Kindling epileptogenesis was induced in rats that received an electrical tetanic stimulation of the Schaffer collateral/commissural fibre pathway in the hippocampus twice daily. Two groups of rats were studied before the appearance of generalized seizures, one group after at least 5 generalized seizures (fully kindled) and one group was investigated at long-term (28 days) after the last seizure. During the initial stages of epileptogenesis, the alpha1A-, alpha1D- and alpha1E-subunit mRNA levels were significantly increased in the different hippocampal subareas in comparison to the levels in control animals. In contrast, alpha1B-subunit gene expression decreased in the CA area and dentate gyrus. No significant change was observed in the alpha1C-I and alpha1C-II expression. At the fully kindled stage, the only significant change was an up-regulation of the alpha1B-subunit mRNA levels in the CA3 area, 24 h after the last seizure. No change in VDCC alpha1-subunit gene expression was found in animals investigated long-term after the establishment of the fully kindled state. Thus, the VDCC alpha1-subunit gene expression is altered in a subclass-specific manner during the early stages of kindling and may play a role in the establishment of a kindled focus, possibly caused by an alteration of the population of VDCCs involved in neurotransmitter release. The absence of long-lasting changes suggests that the maintenance of a kindled focus is not due to persisting alterations in VDCC alpha1 mRNA levels.

Animals↗
PubMed1997

Effect of adrenalectomy in kindled rats.

It has previously been observed that adrenalectomy (ADX) increases the amplitude of transient Ca currents in CA1 hippocampal neurons. In the present study, the effect of ADX on cellular properties of CA1 neurons was investigated under conditions that challenge the CA1 network, i.e. in fully kindled rats, 5-6 weeks after the last kindling session. At this time, adrenally intact kindled rats showed, in comparison to non-kindled controls, a reduced population spike amplitude evoked in the CA1 area by Schaffer/commissural fiber stimulation, an increased amplitude of low-threshold, transient Ca currents and a decrease of the high-threshold, sustained Ca currents. As observed before, ADX in non-kindled rats increased the amplitude of transient Ca currents while sustained Ca currents were not affected. In kindled rats, ADX did not increase the transient Ca current amplitude beyond the level reached by kindling alone. The kindling-induced reduction of the sustained Ca current was partly normalized after ADX. Field responses of kindled rats were not affected by ADX. These data support the view that lack of adrenal hormones does not exacerbate but, to some extent, normalizes the kindling-induced changes in cellular properties and network function of the CA1 area. The hormonal effects on kindling may be reciprocal in nature, since kindling, in turn, was found to affect the stress response and the corticosteroid receptor mRNA expression in the hippocampus.

Adrenal Glands↗
PubMed1996

Corticosteroid actions on the expression of kainate receptor subunit mRNAs in rat hippocampus.

Previous studies have shown that corticosteroid hormones affect kainate-induced excitotoxic processes in the rat hippocampus. In the present study we employed in situ hybridization to examine the effect of adrenalectomy, and subsequent treatment with a low or a high lose of corticosterone on the mRNA levels for kainate receptor subunits in the hippocampus. We observed that adrenalectomy by itself does not affect the expression pattern for the GluR6, GluR7, KAR1 and KAR2 subunits. However, treatment of the adrenalectomized animals with a low dose of corticosterone (3 micrograms/100 g bodyweight) resulted in an enhanced expression of the KAR1, KAR2 and GluR6 subunit mRNAs, when compared to the expression levels in the untreated rats or the sham operated controls. Treatment with a high dose of corticosterone (1 mg/100 g bodyweight) yielded expression levels which were significantly lower than those observed in animals treated with a low corticosterone dose, for the KAR1, KAR2 and GluR7 subunit mRNAs; the levels did not differ from those in untreated rats or in the sham group. We conclude that changes in corticosteroid receptor occupancy, which may occur daily due to circadian or stress-induced variations in the circulating corticosterone level, potentially regulate high affinity kainate receptor activation and the processes in which these receptors are involved.

Adrenal Cortex Hormones↗
PubMed1995

Isozyme specific changes in the expression of protein kinase C isozyme (alpha-zeta) genes in the hippocampus of rats induced by kindling epileptogenesis.

The transcript levels of the protein kinase C (PKC) isoform genes during the development of a kindled epileptogenic focus, elicited by stimulation of Schaffer collateral/commissural fibres in the CA1 area of the rat hippocampus, were compared with the expression levels in control animals using a semi-quantitative in situ hybridization approach. In the hippocampus of control animals, the levels of PKC-alpha-zeta transcripts showed a gene-specific expression pattern and significant differences in expression level were observed between the neurons of CA1, CA3 and fascia dentata. In the early stages of kindling epileptogenesis, i.e. following 6 and 14 afterdischarges, specific changes in the expression levels of PKC-beta, -epsilon, and -zeta but not of PKC-alpha, -gamma, and -delta were found. PKC-beta expression was decreased in CA1, while the PKC-epsilon and -zeta specific hybridization signals were increased in CA1, CA3 and fascia dentata. In fully kindled animals, that had experienced 10 generalized seizures, most expression levels tended to return to control values. One month after the last seizure no significant alterations were encountered. These results indicate an involvement of specific PKC-isoform gene expression in the induction of an epileptogenic focus, but not in the maintenance of the long-lasting kindled state.

Animals↗
PubMed1995

Ischaemia does not alter the editing status at the Q/R site of glutamate receptor-A, -B, -5 and -6 subunit mRNA.

The editing status of mRNA at the Q/R site of the AMPA/kainate type glutamate receptor (GluR) subunits A, B, 5 and 6 modulates the ion selectivity of glutamate receptor operated ion channels. We hypothesized that a long-lasting dysfunction of this editing process after an ischeamic insult may be involved in the process of delayed neuronal cell death. In order to test this hypothesis, the editing status of the GluRs was determined in the hippocampus of rats up to 24 h after 10 min of global ischaemia. No statistical significant differences were found in the hippocampus of animals after ischaemia compared with controls. It is concluded that delayed neuronal cell death is not mediated by a less selective or less efficient mRNA editing process of the different glutamate receptor subunits in the hippocampus.

Animals↗
PubMed1995

Editing status at the Q/R site of glutamate receptor-A, -B, -5 and -6 subunit mRNA in the hippocampal kindling model of epilepsy.

The editing status of mRNA at the Q/R site of the glutamate receptor subtypes -A, -B, -5 and -6 modulates channel conductivity and ion selectivity of glutamate operated ion channels [4,15,26,30]. In order to investigate whether a modification of this editing process may be involved in kindling epileptogenesis, the percentage of edited variant was determined in the hippocampus of kindled rats and compared to the percentage in control animals. In the latter, GluR-A mRNA was detected only in the unedited form (with detection threshold for edited form < 0.7%), whereas GluR-B was completely edited (> 99%). For percentages were not significantly changed in Schaffer collateral/commissural pathway kindled animals that were sacrificed 24 h after the last generalized seizure. It is concluded that the increased sensitivity for the induction of seizures characteristic for Schaffer collateral kindled animals is not related to a less selective or less efficient mRNA editing process of the different glutamate receptor subunits in the hippocampus.

Animals↗
PubMed1995

Expression of GABAA receptor subunit mRNAs in hippocampal pyramidal and granular neurons in the kindling model of epileptogenesis: an in situ hybridization study.

To investigate the molecular changes underlying kindling epileptogenesis in the rat hippocampus, the expression levels of the genes encoding for 13 different gamma-aminobutyric acid type-A receptor (GABAAR) subunits were measured in hippocampal principal neurons using in situ hybridization techniques and semi-quantitative analysis of the autoradiograms. Schaffer collateral-commissural pathway kindled rats were investigated at three different stages of kindling acquisition, at 24 h after the last seizure and at long-term (28 days) after termination of kindling stimulations. Changes were distinct for the different subunits in the three analyzed regions (CA1, CA3, fascia dentata) and also different for the various kindling stages. In all hippocampal areas at the early phases of kindling epileptogenesis, before the appearance of generalized seizures, an increase was found of those transcripts that constituted the majority of the expressed variants in control animals (alpha 1, alpha 2, alpha 4, beta 1, beta 2, beta 3, gamma 2/gamma 2L mRNA). In these stages, the increased levels of different variants in the granular neurons of the fascia dentata were more pronounced when compared to the pattern of changes in pyramidal cells of CA1 and CA3. In fully kindled animals, the expression levels of several subunits returned to control levels, whereas beta 3 and gamma 2/gamma 2L mRNA expression was still significantly enhanced in all areas. At long-term, few changes were encountered. The long-splice variant of gamma 2 was decreased within pyramidal and granular neurons while the total level of gamma 2 mRNA was not different from controls. The increased GABAAR subunit expression in the fascia dentata may underly the reported increased GABAAR ligand binding and the increased GABA mediated inhibition. However, the decreased GABAAR binding and the attenuation of GABAergic inhibition in CA1, could not be explained by a decrement of receptor subunit expression.

Animals↗
PubMed1995

Autoradiographic analysis of [35S]t-butylbicyclophosphorothionate binding in kindled rat hippocampus shows different changes in CA1 area and fascia dentata.

We studied the binding of [35S]t-butylbicyclophosphorothionate to the GABAA receptor-mediated chloride channel in the CA1 area and fascia dentata of control and Schaffer collateral kindled rats, by means of semi-quantitative autoradiography. The [35S]t-butylbicyclophosphorothionate binding was determined at three stages during kindling acquisition: (i) after six afterdischarges, (ii) after 14 afterdischarges and (iii) after the induction of fully kindled seizures. Furthermore, the binding was studied at the long-term stage, 28 days after the last generalized tonic-clonic seizure [Racine R. J. (1972) Electroenceph. clin. Neurophysiol. 32, 281-294]. The binding was investigated at three [35S]t-butylbicyclophosphorothionate concentrations, 4, 47.5 (KD value) and 180 nM (Bmax value). A significant decrease in [35S]t-butylbicyclophosphorothionate binding in the CA1 area (-6 to -20%) and hilar formation (-17 to -37%), in one or more of the three [35S]t-butylbicyclophosphorothionate concentrations tested at the six and 14 afterdischarges and fully kindled stages was observed, but no significant changes at the long-term kindling stage were found. In contrast, the granular and molecular layers of the fascia dentata presented a significant increase in [35S]t-butylbicyclophosphorothionate binding (+15 to +38%) at the 14 afterdischarges, fully kindled and long-term kindled stages.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

N-methyl-D-aspartate and kainate receptor gene expression in hippocampal pyramidal and granular neurons in the kindling model of epileptogenesis.

To investigate the changes underlying kindling epileptogenesis in the rat hippocampus, the levels of the messenger RNAs encoding for the subunits of the N-methyl-D-aspartate-receptor (1, 2A-D) and the kainate-receptor (1, -2, GluR-5, -6, -7) were determined in hippocampal principal neurons using in situ hybridization techniques and semi-quantitative analysis of the autoradiograms. Schaffer collateral-commissural pathway kindled rats were investigated at three different stages of kindling acquisition, always 24 h after the last stimulation. Furthermore, fully kindled rats were studied at long-term (28 days) after termination of kindling stimulations. NR1 messenger RNA levels were slightly decreased in CA1 area of fully kindled animals. In the fascia dentata region, a minor increase of NR2A and NR2B transcripts was found at all stages of kindling acquisition. KA-2 messenger RNA was enhanced in all hippocampal subfields during kindling development. However, none of these changes persisted at long-term after the last seizure and only the low-abundant GluR-7 expression was slightly depressed in the fascia dentata. From our observations we conclude that it is unlikely that alterations in N-methyl-Daspartate or kainate receptor gene expression play an important role in kindling acquisition or maintenance.

Animals↗