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Biomedical subjects

P B Allen

Publications and source records attributed to P B Allen.

At least 19 recordsLinked to original sources
PubMed2006

Distinct roles for spinophilin and neurabin in dopamine-mediated plasticity.

Protein phosphatase 1 plays a major role in the governance of excitatory synaptic activity, and is subject to control via the neuromodulatory actions of dopamine. Mechanisms involved in regulating protein phosphatase 1 activity include interactions with the structurally related cytoskeletal elements spinophilin and neurabin, synaptic scaffolding proteins that are highly enriched in dendritic spines. The requirement for these proteins in dopamine-related neuromodulation was tested using knockout mice. Dopamine D1-mediated regulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor activity was deficient in both striatal and prefrontal cortical neurons from neurabin knockout mice; in spinophilin knockout mice this deficit was manifest only in striatal neurons. At corticostriatal synapses long-term potentiation was deficient in neurabin knockout mice, but not in spinophilin knockout mice, and was rescued by a D1 receptor agonist. In contrast, long-term depression was deficient in spinophilin knockout mice but not in neurabin knockout mice, and was rescued by D2 receptor activation. Spontaneous excitatory post-synaptic current frequency was increased in neurabin knockout mice, but not in spinophilin knockout mice, and this effect was normalized by D2 receptor agonist application. Both knockout strains displayed increased induction of GluR1 Ser(845) phosphorylation in response to D1 receptor stimulation in slices, and also displayed enhanced locomotor activation in response to cocaine administration. These effects could be dissociated from cocaine reward, which was enhanced only in spinophilin knockout mice, and was accompanied by increased immediate early gene induction. These data establish a requirement for synaptic scaffolding in dopamine-mediated responses, and further indicate that spinophilin and neurabin play distinct roles in dopaminergic signal transduction and psychostimulant response.

Animals↗
PubMed2004

Estrogen induces phosphorylation of cyclic AMP response element binding (pCREB) in primary hippocampal cells in a time-dependent manner.

Using hippocampal primary cell cultures at 14 days in vitro (div), we have investigated actions of 17-beta estradiol (E; 10 nM) on the phosphorylation of CREB and on signaling pathways that regulate CREB phosphorylation. After demonstrating that 14 div is optimal for these studies, we examined the time course of E induction of CREB phosphorylation (pCREB) at serine residue 133. The induction of pCREB occurs as early as 1 h following E treatment, presumably via a mechanism involving an E-stimulated signal transduction system, which is sustained for at least 24 h but inhibited by 48 h. The early activity may represent an initial signal required for events leading to phosphorylation of CREB while the sustained signal may lead to CREB-mediated gene expression for cell survival and synapse formation. Furthermore, we examined the pathways for E action preceding pCREB induction by blocking three major kinases (protein kinase; mitogen activated protein kinase, MAPK; and calcium-calmodulin kinase II, CaMKII) upstream of pCREB. We found that E stimulates each pathway at 24 h and that phosphorylation of CREB is dependent on both MAPK and CaMK activities, but less dependent on the Akt pathway. Because CREB has been linked to E induction of excitatory spine synapses, we used a spine marker, spinophilin, to establish E effects on spine formation. Spinophilin expression was up-regulated in response to E and this effect was blocked by an inhibitor of (CaMKII). These studies demonstrate the central role played by CaMKII pathway in the actions of E on both transcriptional regulation and structural reorganization in neurons.

Animals↗
PubMed2004

Estradiol affects spinophilin protein differently in gonadectomized males and females.

Estrogen (E) treatment of ovariectomized animals increases dendritic spines and/or synaptic protein expression in the hippocampus of female rats [J Neurosci 12 (1992) 2549; Endocrinology 142 (2001) 1284; Endocrinol Rev 20 (1999) 279; Annu Rev Pharmacol Toxicol 41 (2001) 569], mice [Proc Natl Acad Sci USA 101 (2004) 2185], rhesus monkeys [Proc Natl Acad Sci USA 98 (2001) 8071; Endocrinology 144 (2003) 4734; J Comp Neurol 465 (2003) 540] and hippocampal cells in vitro [J Neurosci 16 (1996) 4059; Neuroscience 124 (2004) 549]. The role of E in hippocampal synaptic structural plasticity in males is less well understood. In the present study, we have used a recently developed technique to count spinophilin immunogold-reactive (Ir) puncta as well as in situ hybridization to compare E effects on spinophilin-Ir and mRNA in gonadectomized female and male rats 48 h after E treatment. Spinophilin is an established spine marker, which interacts with several proteins (including actin and protein phosphatase 1) that are highly enriched in spines [Proc Natl Acad Sci USA 94 (1997) 9956; Proc Natl Acad Sci USA 97 (2000) 9287]. We report that E exerts sex-specific effects on dendritic spinophilin-labeled spines in the CA1 region: E treatment significantly increased spinophilin-Ir puncta, indicative of spines, in females, but led to a decrease in males. Furthermore, while hippocampal spinophilin mRNA changes could have occurred earlier, spinophilin mRNA levels were unchanged after 48 h of E in both males and females. This suggests the possibility that E regulates spinophilin protein expression and or stability within dendrites via post-transcriptional mechanisms.

Animals↗
PubMed2003

Central and peripheral haemodynamic effects of L-NAME infusion in healthy volunteers.

AIMS: To evaluate the effects of the intravenous administration of the nitric oxide synthesis inhibitor N(g)nitro-L-arginine methyl ester (L-NAME) in healthy volunteers. METHODS: L-NAME (0.25, 0.5 and 0.75 mg/kg over 8 min) was infused in 13 healthy male volunteers. Finally, subjects were infused with either L- or D-arginine. RESULTS: L-NAME resulted in dose-dependent falls in heart rate 60 bpm (55-64 bpm) to 49 bpm (46-52 bpm) (P<0.01) and increased mean arterial pressure 77.0 mmHg (73.2-80.8 mmHg) to 90.0 mmHg (87.1-92.8 mmHg) (P<0.01). The cardiac output was significantly reduced after each L-NAME infusion, and systemic vascular resistance increased linearly over the dosage range. Cardiac stroke volume was significantly reduced only following 0.75 mg/kg/min L-NAME: from 100 ml (91.3-108.7 ml) to 83 ml (74.7-91.4 ml); P<0.01. Forearm blood flow was unchanged at any dosage. L-arginine but not D-arginine infusion reversed the haemodynamic effects of L-NAME. CONCLUSIONS: Contrasting with the profound dose-dependent effects of L-NAME had significant effects on central haemodynamics but no discernible effects on peripheral blood flow.

Dose-Response Relationship, Drug↗
PubMed2001

Optical properties of c-axis oriented superconducting MgB2 films.

Temperature dependent optical conductivities and dc resistivity of c-axis oriented superconducting (T(c) = 39.6 K) MgB2 films (approximately 450 nm) have been measured. The normal state ab-plane optical conductivities can be described by the Drude model with a temperature independent Drude plasma frequency of omega(p,D) = 13 600+/-100 cm(-1) or 1.68+/-0.01 eV. The normal state resistivity is fitted by the Bloch-Grüneisen formula with an electron-phonon coupling constant lambda(tr) = 0.13+/-0.02. The optical conductivity spectra below T(c) of these films suggest that MgB2 is a multigap superconductor.

Journal Article↗
PubMed2001

Is myosin phosphatase regulated in vivo by inhibitor-1? Evidence from inhibitor-1 knockout mice.

1. The Ca(2+) sensitivity of smooth muscle contractility is modulated via regulation of phosphatase activity. Protein phosphatase inhibitor-1 (I-1) is the classic type-1 phosphatase inhibitor, but its presence and role in cAMP-dependent protein kinase (PKA) modulation of smooth muscle is unclear. To address the relevance of I-1 in vivo, we investigated smooth muscle function in a mouse model lacking the I-1 protein (I-1((-/-)) mice). 2. Significant amounts of I-1 protein were detected in the wild-type (WT) mouse aorta and could be phosphorylated by PKA, as indicated by (32)P-labelled aortic extracts from WT mice. 3. Despite the significant presence of I-1 in WT aorta, phenylephrine and KCl concentration- isometric force relations in the presence or absence of the PKA pathway activator isoproterenol (isoprenaline) were unchanged compared to I-1((-/-)) aorta. cGMP-dependent protein kinase (PKG) relaxation pathways were also not different. Consistent with these findings, dephosphorylation rates of the 20 kDa myosin light chains (MLC(20)), measured in aortic extracts, were nearly identical between WT and I-1((-/-)) mice. 4. In the portal vein, I-1 protein ablation was associated with a significant (P < 0.05) rightward shift in the EC(50) of isoproterenol relaxation (EC(50) = 10.4 +/- 1.4 nM) compared to the WT value (EC(50) = 3.5 +/- 0.2 nM). Contraction in response to acetylcholine as well as Ca(2+) sensitivity were similar between WT and I-1((-/-)) aorta. 5. Despite the prevalence of I-1 and its activation by PKA in the aorta, I-1 does not appear to play a significant role in contractile or relaxant responses to any pharmacomechanical or electromechanical agonists used. I-1 may play a role as a fine-tuning mechanism involved in regulating portal vein responsiveness to beta-adrenergic agonists.

Acetylcholine↗
PubMed2001

Protein phosphatase 1 regulation by inhibitors and targeting subunits.

Regulation of protein phosphatase 1 (PP1) by protein inhibitors and targeting subunits has been previously studied through the use of recombinant protein expressed in Escherichia coli. This preparation is limited by several key differences in its properties compared with native PP1. In the present study, we have analyzed recombinant PP1 expressed in Sf9 insect cells using baculovirus. Sf9 PP1 exhibited properties identical to those of native PP1, with respect to regulation by metals, inhibitor proteins, and targeting subunits, and failure to dephosphorylate a phosphotyrosine-containing substrate or phospho-DARPP-32 (Dopamine and cAMP-regulated phosphoprotein, M(r) 32,000). Mutations at Y272 in the beta12/beta13 loop resulted in a loss of activity and reduced the sensitivity to thiophospho-DARPP-32 and inhibitor-2. Mutations of Y272 also increased the relative activity toward a phosphotyrosine-containing substrate or phospho-DARPP-32. Mutation of acidic groove residues caused no change in sensitivity to thiophospho-DARPP-32 or inhibitor-2, but one mutant (E252A:D253A:E256R) exhibited an increased K(m) for phosphorylase a. Several PP1/PP2A chimeras were prepared in which C-terminal sequences of PP2A were substituted into PP1. Replacement of residues 274-330 of PP1 with the corresponding region of PP2A resulted in a large loss of sensitivity to thiophospho-DARPP-32 and inhibitor-2, and also resulted in a loss of interaction with the targeting subunits, spinophilin and PP1 nuclear targeting subunit (PNUTS). More limited alterations in residues in beta12, beta13, and beta14 strands highlighted a key role for M290 and C291 in the interaction of PP1 with thiophospho-DARPP-32, but not inhibitor-2.

Amino Acid Sequence↗
PubMed2001

Antipsychotic treatment induces alterations in dendrite- and spine-associated proteins in dopamine-rich areas of the primate cerebral cortex.

BACKGROUND: Mounting evidence indicates that long-term treatment with antipsychotic medications can alter the morphology and connectivity of cellular processes in the cerebral cortex. The cytoskeleton plays an essential role in the maintenance of cellular morphology and is subject to regulation by intracellular pathways associated with neurotransmitter receptors targeted by antipsychotic drugs. METHODS: We have examined whether chronic treatment with the antipsychotic drug haloperidol interferes with phosphorylation state and tissue levels of a major dendritic cytoskeleton-stabilizing agent, microtubule-associated protein 2 (MAP2), as well as levels of the dendritic spine-associated protein spinophilin and the synaptic vesicle-associated protein synaptophysin in various regions of the cerebral cortex of rhesus monkeys. RESULTS: Among the cortical areas examined, the prefrontal, orbital, cingulate, motor, and entorhinal cortices displayed significant decreases in levels of spinophilin, and with the exception of the motor cortex, each of these regions also exhibited increases in the phosphorylation of MAP2. No changes were observed in either spinophilin levels or MAP2 phosphorylation in the primary visual cortex. Also, no statistically significant changes were found in tissue levels of MAP2 or synaptophysin in any of the cortical regions examined. CONCLUSIONS: Our findings demonstrate that long-term haloperidol exposure alters neuronal cytoskeleton- and spine-associated proteins, particularly in dopamine-rich regions of the primate cerebral cortex, many of which have been implicated in the psychopathology of schizophrenia. The ability of haloperidol to regulate cytoskeletal proteins should be considered in evaluating the mechanisms of both its palliative actions and its side effects.

Animals↗
PubMed2001

Novel target sites for estrogen action in the dorsal hippocampus: an examination of synaptic proteins.

Structural studies have shown that estrogens increase dendritic spine number in the dorsal CA1 field of rat hippocampus using Golgi impregnation as well as the number of dorsal CA1 synapses visualized via electron microscopy. The present study was carried out to further these findings by examining changes in the levels of pre- and postsynaptic proteins using radioimmunocytochemistry (RICC). In this study, 2 days of estradiol-benzoate treatment produced significant and comparable increases in synaptophysin, syntaxin, and spinophilin immunoreactivity (IR) in the CA1 region of the dorsal hippocampus of ovariectomized female rats. For spinophilin, IR was also increased in the hilar region of the dentate gyrus as well as CA3. In all cases, the nonsteroidal estrogen antagonist CI628, which has been previously shown to block spine formation, inhibited the effects of estrogen. However, these protein differences were not detected in whole hippocampus using Western blots. These findings add to a growing body of evidence that estrogens increase synapses in the CA1 region of hippocampus along with changes in previously unidentified sites. These results also suggest that RICC is a rapid and sensitive method for examining molecular changes in synaptic profiles in anatomically distinct brain regions.

Animals↗
PubMed2001

Nitric oxide modulation of blood vessel tone identified by arterial waveform analysis.

Traditionally, nitric oxide-mediated alteration in blood vessel tone has been inferred from changes in flow in response to physical and pharmacological interventions using plethysmographic or ultrasonic techniques. We hypothesized that alteration in pulsatile arterial function may represent a more sensitive measure to detect and monitor nitric oxide-mediated modulation of arterial smooth muscle tone. Healthy male volunteers (n = 15) had radial artery pressure pulse waveforms recorded using a calibrated tonometer device. A computer-based assessment of the diastolic pressure decay was employed to quantify changes in arterial waveform morphology in terms of altered pulsatile (arterial compliance) and steady-state (peripheral resistance) haemodynamics. N(G)-nitro-L-arginine methyl ester (L-NAME), a stereospecific inhibitor of nitric oxide synthesis, was infused intravenously in incrementally increasing doses of 0.25, 0.5 and 0.75 mg/kg for 8 min each. Subjects then received either L-arginine or D-arginine (200 mg/kg over 15 min) intravenously in a blinded fashion. On a separate day, subjects had radial artery pressure pulse waveforms recorded before and after the sublingual administration of glyceryl trinitrate, an exogenous donor of nitric oxide. Cardiac output and heart rate decreased and mean arterial blood pressure increased significantly (P < 0.01 for all) in response to the incremental intravenous infusion of L-NAME. Small artery compliance decreased, whereas systemic vascular resistance increased in response to nitric oxide synthesis inhibition (P < 0.01 for both). The intravenous infusion of L-arginine restored the pulsatile and steady-state haemodynamic parameters to pre-treatment values, whereas D-arginine had no effect. Sublingual glyceryl trinitrate decreased systemic vascular resistance by 11%, whereas large artery- and small artery-compliance increased by 25% and 44% respectively. Pressure pulse contour analysis represents a sensitive and convenient technique capable of tracking changes in the pulsatile function of arteries accompanying nitric oxide-mediated alteration in arterial smooth muscle tone.

Adult↗
PubMed2000

Spinophilin regulates the formation and function of dendritic spines.

Spinophilin, a protein that interacts with actin and protein phosphatase-1, is highly enriched in dendritic spines. Here, through the use of spinophilin knockout mice, we provide evidence that spinophilin modulates both glutamatergic synaptic transmission and dendritic morphology. The ability of protein phosphatase-1 to regulate the activity of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptors was reduced in spinophilin knockout mice. Consistent with altered glutamatergic transmission, spinophilin-deficient mice showed reduced long-term depression and exhibited resistance to kainate-induced seizures and neuronal apoptosis. In addition, deletion of the spinophilin gene caused a marked increase in spine density during development in vivo as well as altered filopodial formation in cultured neurons. In conclusion, spinophilin appears to be required for the regulation of the properties of dendritic spines.

Animals↗
PubMed2000

Regulation of phosphorylation of the GluR1 AMPA receptor in the neostriatum by dopamine and psychostimulants in vivo.

The activation of cAMP-dependent protein kinase regulates the physiological activity of AMPA-type glutamate receptors. In this study, phosphorylation of the AMPA receptor subunit GluR1 at Ser(845) was increased in neostriatal slices by activation of D1-type dopamine receptors and by inhibitors of protein phosphatase 1/protein phosphatase 2A. In contrast, Ser(831), a residue which, when phosphorylated by protein kinase C or calcium/calmodulin-dependent kinase II, increases AMPA receptor channel conductance, was unaffected by either D1 or D2 receptor agonists in neostriatal slices. The phosphorylation of Ser(845), but not Ser(831), was strongly increased in neostriatum in vivo in response to the psychostimulants cocaine and methamphetamine. The effects of dopamine and psychostimulants on the phosphorylation of GluR1 were attenuated in dopamine and cAMP-regulated phosphoprotein M(r) 32 kDa (DARPP-32) knock-out mice. These results identify DARPP-32 and AMPA-type glutamate receptors as likely essential cellular effectors for psychostimulant actions.

Animals↗
PubMed2000

Protein phosphatase-1 regulation in the induction of long-term potentiation: heterogeneous molecular mechanisms.

Protein phosphatase inhibitor-1 (I-1) has been proposed as a regulatory element in the signal transduction cascade that couples postsynaptic calcium influx to long-term changes in synaptic strength. We have evaluated this model using mice lacking I-1. Recordings made in slices prepared from mutant animals and also in anesthetized mutant animals indicated that long-term potentiation (LTP) is deficient at perforant path-dentate granule cell synapses. In vitro, this deficit was restricted to synapses of the lateral perforant path. LTP at Schaffer collateral-CA1 pyramidal cell synapses remained normal. Thus, protein phosphatase-1-mediated regulation of NMDA receptor-dependent synaptic plasticity involves heterogeneous molecular mechanisms, in both different dendritic subregions and different neuronal subtypes. Examination of the performance of I-1 mutants in spatial learning tests indicated that intact LTP at lateral perforant path-granule cell synapses is either redundant or is not involved in this form of learning.

Animals↗
PubMed2000

Requirement for DARPP-32 in progesterone-facilitated sexual receptivity in female rats and mice.

DARPP-32, a dopamine- and adenosine 3',5'-monophosphate (cAMP)-regulated phosphoprotein (32 kilodaltons in size), is an obligate intermediate in progesterone (P)-facilitated sexual receptivity in female rats and mice. The facilitative effect of P on sexual receptivity in female rats was blocked by antisense oligonucleotides to DARPP-32. Homozygous mice carrying a null mutation for the DARPP-32 gene exhibited minimal levels of P-facilitated sexual receptivity when compared to their wild-type littermates. P significantly increased hypothalamic cAMP levels and cAMP-dependent protein kinase activity. These increases were not inhibited by a D1 subclass dopamine receptor antagonist. P also enhanced phosphorylation of DARPP-32 on threonine 34 in the hypothalamus of mice. DARPP-32 activation is thus an obligatory step in progestin receptor regulation of sexual receptivity in rats and mice.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed2000

Dopamine D(1) receptor-induced gene transcription is modulated by DARPP-32.

The role of the dopamine- and cyclic AMP-regulated phosphoprotein of M(r) 32,000 (DARPP-32) in dopaminergic regulation of gene transcription in striatum and globus pallidus was examined. Mice with targeted disruption of the gene encoding DARPP-32, its homologue, inhibitor-1, or both, were used. Pharmacological characterization showed that mutant mice had normal basal levels of dopamine D(1) and D(2) receptors and adenosine A(2A) receptors. Basal expression levels of the striatonigral-specific neuropeptides substance P and prodynorphin and the immediate early genes c-fos and NGFI-A were also unaltered in mutant mice. A full D(1) receptor agonist, SKF 82958, up-regulated the expression of these neuropeptides and immediate early genes significantly more in wild-type mice than in mice lacking DARPP-32. Moreover, the additive stimulation of SKF 82958 and quinelorane, a D(2) receptor agonist, on c-fos mRNA in globus pallidus was significantly decreased in DARPP-32 and DARPP-32/I-1 knockout mice. No changes in dopamine receptor-induced gene expression were found in I-1 knockout mice. These results demonstrate an important involvement of DARPP-32 in dopamine receptor-mediated regulation of gene expression both in striatal neurons, which are enriched in DARPP-32, and in pallidal neurons, which do not contain DARPP-32.

Animals↗
PubMed2000

Howard H. Hepburn and the development of skull tongs for cervical spine traction.

The first use of skull tongs for cervical spine traction is credited primarily to W.G. Crutchfield. In 1933, Crutchfield described his application of extension tongs to the calvaria of a 23-year-old woman with a traumatic C2-C3 fracture. Less recognized are the contributions of Howard H. Hepburn, who designed skull tongs for cervical spine traction at the University of Alberta several years before Crutchfield's first case. Hepburn was the first neurosurgeon at the University of Alberta in Edmonton. On the basis of his experience treating wounded soldiers in World War I, he developed the hypothesis that traction would promote healing in cervical spine injuries. Hepburn designed skull extension tongs that were modeled on common ice tongs, and he used an automobile inner tube as an elastic to keep the tongs firmly applied to the patient's head. These tongs were first used in the mid-1920s, and by 1930 they were applied routinely. Crutchfield's 1933 report refers to the application of "Edmonton extension tongs." This suggests that he was at least indirectly aware of Hepburn's work, although how this information reached him is not entirely clear. Hepburn attended a meeting of the British Medical Society in 1930, and he is thought to have discussed his tongs during the conference. Hepburn's work has received some attention previously; his original tongs were included in a 1973 Smithsonian Institute exhibit on cervical spine traction as an example of an early cranial traction device. However, his contributions are underappreciated in the neurosurgical community and deserve wider recognition.

Canada↗