The Journal of comparative neurology, Jan 26, 1998
Alternative splicing has been shown to occur at the metabotropic glutamate receptor 1 (mGluR1) ge... more Alternative splicing has been shown to occur at the metabotropic glutamate receptor 1 (mGluR1) gene. Three main isoforms that differ in their carboxy-termini have been described so far and named mGluR1alpha, mGluR1beta and mGluR1c. These variants when expressed in recombinant systems all activate phospholipase C, although the [Ca2+] signals generated have different kinetics. Tissue distribution studies of specific mGluR1 splice variants are limited to the mGluR1alpha isoform. In the present work, we examined the localization of mGluR1beta in the adult rat and mouse forebrain by using a specific antipeptide antibody. Furthermore, the mGluR1beta immunostaining was compared with that obtained with antibodies specific for mGluR1alpha or with a pan-mGluR1 antibody which recognizes all isoforms. mGluR1beta-like immunoreactivity (LI) was found confined to the neuropil and neuronal perikarya and appeared discretely distributed in the rodent forebrain. Differential cellular distribution betw...
Optical recording of neuronal activities using voltage-sensitive dyes (VSDs) is a useful method f... more Optical recording of neuronal activities using voltage-sensitive dyes (VSDs) is a useful method for simultaneous multi-site recording. However, it has been rather difficult to distinguish optical signals from individual, identified cells. We applied the optical recording technique using a high-speed charge coupled device (CCD) imaging system to a teleost thalamic nucleus, corpus glomerulosum (CG) which has a well-defined histological organization and large postsynaptic dendrites. Patch-like dye (di-4-ANEPPS) signals were observed in the dendritic layer of the CG in response to afferent nerve stimulations. These responses were completely blocked by an alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionate (AMPA) receptor antagonist, did not propagate, and the size of the patches were close to that of a single dendritic tip of the 'large cell'. Thus, we found that these patch-like VSD signals most likely represent postsynaptic potentials at individual dendritic tips of the large cells.
World journal of biological chemistry, Jan 26, 2010
The cerebellum expresses one of the highest levels of the plasma membrane Ca(2+) ATPase, isoform ... more The cerebellum expresses one of the highest levels of the plasma membrane Ca(2+) ATPase, isoform 2 in the mammalian brain. This highly efficient plasma membrane calcium transporter protein is enriched within the main output neurons of the cerebellar cortex; i.e. the Purkinje neurons (PNs). Here we review recent evidence, including electrophysiological and calcium imaging approaches using the plasma membrane calcium ATPase 2 (PMCA2) knockout mouse, to show that PMCA2 is critical for the physiological control of calcium at cerebellar synapses and cerebellar dependent behaviour. These studies have also revealed that deletion of PMCA2 throughout cerebellar development in the PMCA2 knockout mouse leads to permanent signalling and morphological alterations in the PN dendrites. Whilst these findings highlight the importance of PMCA2 during cerebellar synapse function and development, they also reveal some limitations in the use of the PMCA2 knockout mouse and the need for additional experi...
Genetic deletion of the plasma membrane calcium ATPase type 2 (PMCA2), a calcium transporter prot... more Genetic deletion of the plasma membrane calcium ATPase type 2 (PMCA2), a calcium transporter protein, is associated with an overtly ataxic phenotype in mice. PMCA2 is expressed at high levels in cerebellar Purkinje neurons (PNs) where functional integrity is essential for normal cerebellar function. Indeed, loss of PN function accompanies cerebellar ataxia in humans and mouse models. In the ataxic PMCA2 knockout (PMCA2-/-) mouse the ability of the PNs to control their cytosolic calcium levels was severely impaired; basal calcium levels were high and calcium recovery kinetics slow. Whole cell patch clamp recordings from PMCA2-/- PNs revealed that they possessed hyperpolarised membrane potentials, reduced frequency and increased irregularity of spontaneous action potential firing, curtailed complex spikes and sustained calcium-dependent outward K+ currents. We propose that these alterations limit pathological excursions in PN cytosolic calcium as an aid to survival but that they are i...
The effects of brief (2-4 min) hypoxia on presumed dopaminergic "principal" neurons of ... more The effects of brief (2-4 min) hypoxia on presumed dopaminergic "principal" neurons of the rat ventral mesencephalon were investigated by using either intracellular or whole cell patch-clamp recordings in in vitro conditions. Under single-electrode voltage clamp, with sharp microelectrode (Vh -60 mV), a brief hypoxia caused an outward current (hypoOUT) of 110.2 +/- 15.2 (SE) pA (n = 18), which was followed by a posthypoxic outward current (posthypoOUT) of 149.6 +/- 10.6 pA (n = 18). Although the hypoOUT reversed at -83.7 +/- 3.8 mV (n = 18), the posthypoOUT did not reverse. The K+ATP-blocking sulphonylureas tolbutamide (100 microM) and glibenclamide (30 microM), significantly reduced the peak of the hypoOUT by 47.6 +/- 7.7% (n = 16) and 54.18 +/- 7.5% (n = 3), respectively. In contrast, they did not affect the posthypoOUT. Extracellular barium (300 microM to 1 mM) almost abolished the hypoOUT, leaving the posthypoOUT unchanged. The large K+ channel blocker charybdotoxin (1...
Slices were prepared from septal and hippocampal tissue and co-cultured for periods up to one mon... more Slices were prepared from septal and hippocampal tissue and co-cultured for periods up to one month. The presence of cholinergic neurons within the septal slices was demonstrated by histochemical staining techniques for acetylcholinesterase or by Golgi-like immunoperoxidase techniques with antibodies raised against the enzyme choline acetyltransferase. Cholinergic fibers originating in the septal explants started to grow radially in all directions. By day 7, the first fibers were seen to reach their target, but maximal hippocampal ingrowth occurred between day 8 and 14 in vitro. Only those fibers reaching the target were maintained, whereas cholinergic fibers growing in other directions degenerated. Electrophysiological studies showed that cholinergic fibers established functional cholinergic connections with hippocampal pyramidal cells. As a result of septal stimulation, two different potassium currents were inhibited in pyramidal cells: a calcium-independent current, IM, and a cal...
Glucose deprivation (GD) results in a hyperpolarization by turning on a potassium conductance (gK... more Glucose deprivation (GD) results in a hyperpolarization by turning on a potassium conductance (gK,GD) in hippocampal CA3 pyramidal cells. We used combined intracellular and microfluorometric recording techniques to evaluate whether gK,GD is activated by a rise in the concentration of intracellular calcium ([Ca2+]i). We found that the activation of gK,GD is only followed, but not preceded by a rise in [Ca2+]i. Furthermore, gK,GD is not blocked by the sulfonylurea glibenclamide, a blocker of ATP-regulated potassium conductance. We conclude that activation of gK,GD does not simply reflect breakdown of the calcium of ATP homeostasis, but on the contrary might represent an active restoring mechanism which delays the pathological consequences of sustained glucose deficiency.
The Journal of comparative neurology, Jan 26, 1998
Alternative splicing has been shown to occur at the metabotropic glutamate receptor 1 (mGluR1) ge... more Alternative splicing has been shown to occur at the metabotropic glutamate receptor 1 (mGluR1) gene. Three main isoforms that differ in their carboxy-termini have been described so far and named mGluR1alpha, mGluR1beta and mGluR1c. These variants when expressed in recombinant systems all activate phospholipase C, although the [Ca2+] signals generated have different kinetics. Tissue distribution studies of specific mGluR1 splice variants are limited to the mGluR1alpha isoform. In the present work, we examined the localization of mGluR1beta in the adult rat and mouse forebrain by using a specific antipeptide antibody. Furthermore, the mGluR1beta immunostaining was compared with that obtained with antibodies specific for mGluR1alpha or with a pan-mGluR1 antibody which recognizes all isoforms. mGluR1beta-like immunoreactivity (LI) was found confined to the neuropil and neuronal perikarya and appeared discretely distributed in the rodent forebrain. Differential cellular distribution betw...
Optical recording of neuronal activities using voltage-sensitive dyes (VSDs) is a useful method f... more Optical recording of neuronal activities using voltage-sensitive dyes (VSDs) is a useful method for simultaneous multi-site recording. However, it has been rather difficult to distinguish optical signals from individual, identified cells. We applied the optical recording technique using a high-speed charge coupled device (CCD) imaging system to a teleost thalamic nucleus, corpus glomerulosum (CG) which has a well-defined histological organization and large postsynaptic dendrites. Patch-like dye (di-4-ANEPPS) signals were observed in the dendritic layer of the CG in response to afferent nerve stimulations. These responses were completely blocked by an alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionate (AMPA) receptor antagonist, did not propagate, and the size of the patches were close to that of a single dendritic tip of the 'large cell'. Thus, we found that these patch-like VSD signals most likely represent postsynaptic potentials at individual dendritic tips of the large cells.
World journal of biological chemistry, Jan 26, 2010
The cerebellum expresses one of the highest levels of the plasma membrane Ca(2+) ATPase, isoform ... more The cerebellum expresses one of the highest levels of the plasma membrane Ca(2+) ATPase, isoform 2 in the mammalian brain. This highly efficient plasma membrane calcium transporter protein is enriched within the main output neurons of the cerebellar cortex; i.e. the Purkinje neurons (PNs). Here we review recent evidence, including electrophysiological and calcium imaging approaches using the plasma membrane calcium ATPase 2 (PMCA2) knockout mouse, to show that PMCA2 is critical for the physiological control of calcium at cerebellar synapses and cerebellar dependent behaviour. These studies have also revealed that deletion of PMCA2 throughout cerebellar development in the PMCA2 knockout mouse leads to permanent signalling and morphological alterations in the PN dendrites. Whilst these findings highlight the importance of PMCA2 during cerebellar synapse function and development, they also reveal some limitations in the use of the PMCA2 knockout mouse and the need for additional experi...
Genetic deletion of the plasma membrane calcium ATPase type 2 (PMCA2), a calcium transporter prot... more Genetic deletion of the plasma membrane calcium ATPase type 2 (PMCA2), a calcium transporter protein, is associated with an overtly ataxic phenotype in mice. PMCA2 is expressed at high levels in cerebellar Purkinje neurons (PNs) where functional integrity is essential for normal cerebellar function. Indeed, loss of PN function accompanies cerebellar ataxia in humans and mouse models. In the ataxic PMCA2 knockout (PMCA2-/-) mouse the ability of the PNs to control their cytosolic calcium levels was severely impaired; basal calcium levels were high and calcium recovery kinetics slow. Whole cell patch clamp recordings from PMCA2-/- PNs revealed that they possessed hyperpolarised membrane potentials, reduced frequency and increased irregularity of spontaneous action potential firing, curtailed complex spikes and sustained calcium-dependent outward K+ currents. We propose that these alterations limit pathological excursions in PN cytosolic calcium as an aid to survival but that they are i...
The effects of brief (2-4 min) hypoxia on presumed dopaminergic "principal" neurons of ... more The effects of brief (2-4 min) hypoxia on presumed dopaminergic "principal" neurons of the rat ventral mesencephalon were investigated by using either intracellular or whole cell patch-clamp recordings in in vitro conditions. Under single-electrode voltage clamp, with sharp microelectrode (Vh -60 mV), a brief hypoxia caused an outward current (hypoOUT) of 110.2 +/- 15.2 (SE) pA (n = 18), which was followed by a posthypoxic outward current (posthypoOUT) of 149.6 +/- 10.6 pA (n = 18). Although the hypoOUT reversed at -83.7 +/- 3.8 mV (n = 18), the posthypoOUT did not reverse. The K+ATP-blocking sulphonylureas tolbutamide (100 microM) and glibenclamide (30 microM), significantly reduced the peak of the hypoOUT by 47.6 +/- 7.7% (n = 16) and 54.18 +/- 7.5% (n = 3), respectively. In contrast, they did not affect the posthypoOUT. Extracellular barium (300 microM to 1 mM) almost abolished the hypoOUT, leaving the posthypoOUT unchanged. The large K+ channel blocker charybdotoxin (1...
Slices were prepared from septal and hippocampal tissue and co-cultured for periods up to one mon... more Slices were prepared from septal and hippocampal tissue and co-cultured for periods up to one month. The presence of cholinergic neurons within the septal slices was demonstrated by histochemical staining techniques for acetylcholinesterase or by Golgi-like immunoperoxidase techniques with antibodies raised against the enzyme choline acetyltransferase. Cholinergic fibers originating in the septal explants started to grow radially in all directions. By day 7, the first fibers were seen to reach their target, but maximal hippocampal ingrowth occurred between day 8 and 14 in vitro. Only those fibers reaching the target were maintained, whereas cholinergic fibers growing in other directions degenerated. Electrophysiological studies showed that cholinergic fibers established functional cholinergic connections with hippocampal pyramidal cells. As a result of septal stimulation, two different potassium currents were inhibited in pyramidal cells: a calcium-independent current, IM, and a cal...
Glucose deprivation (GD) results in a hyperpolarization by turning on a potassium conductance (gK... more Glucose deprivation (GD) results in a hyperpolarization by turning on a potassium conductance (gK,GD) in hippocampal CA3 pyramidal cells. We used combined intracellular and microfluorometric recording techniques to evaluate whether gK,GD is activated by a rise in the concentration of intracellular calcium ([Ca2+]i). We found that the activation of gK,GD is only followed, but not preceded by a rise in [Ca2+]i. Furthermore, gK,GD is not blocked by the sulfonylurea glibenclamide, a blocker of ATP-regulated potassium conductance. We conclude that activation of gK,GD does not simply reflect breakdown of the calcium of ATP homeostasis, but on the contrary might represent an active restoring mechanism which delays the pathological consequences of sustained glucose deficiency.
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