Blockade of AMPA Receptors and Volatile Anesthetics

Background The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) subtype of glutamate receptor mediates fast excitatory neurotransmission in the central nervous system. Many general anesthetics inhibit AMPA receptors in vitro; however, it is not certain if this inhibition contributes to the behavioral properties of these drugs. AMPA receptors lacking the GluR2 subunit are resistant to blockade by barbiturates in vitro. Paradoxically, GluR2 null mutant (-/-) mice are more sensitive to barbiturate-induced loss of the righting reflex (LORR) compared with wild-type (+/+) littermates. To determine if interactions between anesthetics and AMPA receptors account for the increased sensitivity of (-/-) mice, the effects of volatile anesthetics that do not directly inhibit AMPA receptors were examined. Methods Isoflurane, halothane, desflurane, or sevoflurane were administered to (-/-) and (+/+) littermate controls. Anesthetic requirements for LORR, movement to tail clamp (minimum alveolar concentration [MAC]), and hind-paw withdrawal latency (HPWL) were determined. Electrophysiologic methods examined the inhibition of AMPA receptors by isoflurane and halothane. Results Anesthetic requirements for LORR and HPWL were decreased, whereas MAC values were unchanged in (-/-) mice. Isoflurane and halothane caused minimal inhibition of AMPA receptors at clinically relevant concentrations. Conclusions Direct blockade of AMPA receptors did not account for the increased sensitivity to volatile anesthetics in GluR2 null mutant mice for HPWL or LORR. Thus, the deficiency of GluR2-containing AMPA receptors increases the sensitivity of neuronal circuitry mediating these end points, but not MAC. GluR2-containing receptors do not contribute appreciably to MAC in this mouse model. These results illustrate the difficulties in attributing behavioral responses to drug-receptor interactions in genetically engineered animals.

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Null Mutation in Transforming Growth Factor β1 Disrupts Ovarian Function and Causes Oocyte Incompetence and Early Embryo Arrest

Endocrinology ◽

10.1210/en.2005-1189 ◽

2006 ◽

Vol 147 (2) ◽

pp. 835-845 ◽

Cited By ~ 51

Author(s):

Wendy V. Ingman ◽

Rebecca L. Robker ◽

Karen Woittiez ◽

Sarah A. Robertson

Keyword(s):

Transforming Growth Factor ◽

Ovarian Function ◽

Mutant Mice ◽

Reproductive Capacity ◽

Null Mutation ◽

Null Mutant ◽

Critical Determinant ◽

Serum Progesterone ◽

Null Mutant Mice

TGFβ1 is implicated in regulation of ovarian function and the events of early pregnancy. We have investigated the effect of null mutation in the Tgfβ1 gene on reproductive function in female mice. The reproductive capacity of TGFβ1 null mutant females was severely impaired, leading to almost complete infertility. Onset of sexual maturity was delayed, after which ovarian function was disrupted, with extended ovarian cycles, irregular ovulation, and a 40% reduction in oocytes ovulated. Serum FSH and estrogen content were normal, but TGFβ1 null mutant mice failed to display the characteristic proestrus surge in circulating LH. Ovarian hyperstimulation with exogenous gonadotropins elicited normal ovulation rates in TGFβ1 null mutant mice. After mating with wild-type stud males, serum progesterone content was reduced by 75% associated with altered ovarian expression of mRNAs encoding steroidogenic enzymes 3β-hydroxysteroid dehydrogenase-1 and P450 17 α-hydroxylase/C17–20-lyase. Embryos recovered from TGFβ1 null mutant females were developmentally arrested in the morula stage and rarely progressed to blastocysts. Attempts to rescue embryos by exogenous progesterone administration and in vitro culture were unsuccessful, and in vitro fertilization and culture experiments demonstrated that impaired development is unlikely to result from lack of maternal tract TGFβ1. We conclude that embryo arrest is due to developmental incompetence in oocytes developed in a TGFβ1-deficient follicular environment. This study demonstrates that TGFβ1 is a critical determinant of normal ovarian function, operating through regulation of LH activity and generation of oocytes competent for embryonic development and successful initiation of pregnancy.

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Neurons Expressing the Highest Levels of γ-Synuclein Are Unaffected by Targeted Inactivation of the Gene

Molecular and Cellular Biology ◽

10.1128/mcb.23.22.8233-8245.2003 ◽

2003 ◽

Vol 23 (22) ◽

pp. 8233-8245 ◽

Cited By ~ 48

Author(s):

Natalia Ninkina ◽

Katerina Papachroni ◽

Darren C. Robertson ◽

Oliver Schmidt ◽

Liz Delaney ◽

...

Keyword(s):

Es Cells ◽

Culture Conditions ◽

Sensory Function ◽

Mutant Mice ◽

Null Mutant ◽

Wild Type ◽

Complete Inactivation ◽

Null Mutant Mice

ABSTRACT Homologous recombination in ES cells was employed to generate mice with targeted deletion of the first three exons of the γ-synuclein gene. Complete inactivation of gene expression in null mutant mice was confirmed on the mRNA and protein levels. Null mutant mice are viable, are fertile, and do not display evident phenotypical abnormalities. The effects of γ-synuclein deficiency on motor and peripheral sensory neurons were studied by various methods in vivo and in vitro. These two types of neurons were selected because they both express high levels of γ-synuclein from the early stages of mouse embryonic development but later in the development they display different patterns of intracellular compartmentalization of the protein. We found no difference in the number of neurons between wild-type and null mutant animals in several brain stem motor nuclei, in lumbar dorsal root ganglia, and in the trigeminal ganglion. The survival of γ-synuclein-deficient trigeminal neurons in various culture conditions was not different from that of wild-type neurons. There was no difference in the numbers of myelinated and nonmyelinated fibers in the saphenous nerves of these animals, and sensory reflex thresholds were also intact in γ-synuclein null mutant mice. Nerve injury led to similar changes in sensory function in wild-type and mutant mice. Taken together, our data suggest that like α-synuclein, γ-synuclein is dispensable for the development and function of the nervous system.

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Blockade of Glutamate Receptors and Barbiturate Anesthesia

Anesthesiology ◽

10.1097/00000542-199911000-00025 ◽

1999 ◽

Vol 91 (5) ◽

pp. 1329-1329 ◽

Cited By ~ 35

Author(s):

Daisy T. Joo ◽

Zhigang Xiong ◽

John F. MacDonald ◽

Zhengping Jia ◽

John Roder ◽

...

Keyword(s):

Ampa Receptor ◽

Ampa Receptors ◽

Sleep Time ◽

Mutant Mice ◽

Gamma Aminobutyric Acid ◽

Null Mutant ◽

Wild Type ◽

Gaba A ◽

Receptor Complexes ◽

Null Mutant Mice

Background Barbiturates enhance gamma-aminobutyric acid type A (GABA(A)) receptor function and also inhibit the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) subtype of glutamate receptor. The relative contribution of these actions to the behavioral properties of barbiturates is not certain. Because AMPA receptor complexes that lack the GluR2 subunit are relatively insensitive to pentobarbital inhibition, GluR2 null mutant mice provide a novel tool to investigate the importance of AMPA receptor inhibition to the anesthetic effects of barbiturates. Methods GluR2 null allele (-/-), heterozygous (+/-), and wild-type (+/+) mice were injected with pentobarbital (30 and 35 mg/kg intraperitoneally). Sensitivity to anesthetics was assessed by measuring the latency to loss of righting reflex, sleep time, and the loss of corneal, pineal, and toe-pinch withdrawal reflexes. In addition, patch-clamp recordings of acutely dissociated CA1 hippocampal pyramidal neurons from (-/-) and (+/+) mice were undertaken to investigate the effects of barbiturates on kainate-activated AMPA receptors and GABA-activated GABA(A) receptors. Results Behavioral tests indicate that sensitivity to pentobarbital was increased in (-/-) mice. In contrast, AMPA receptors from (-/-) neurons were less sensitive to inhibition by pentobarbital (concentrations that produced 50% of the maximal inhibition [IC50], 301 vs. 51 microM), thiopental (IC50, 153 vs. 34 microM), and phenobarbital (IC50, 930 vs. 205 microM) compared with wild-type controls, respectively. In addition, the potency of kainate was greater in (-/-) neurons, whereas no differences were observed for the potentiation of GABA(A) receptors by pentobarbital. Conclusions The GluR2 null mutant mice were more sensitive to pentobarbital anesthesia despite a reduced sensitivity of GluR2-deficient AMPA receptors to barbiturate blockade. Our results indicate that the inhibition of AMPA receptors does not correlate with the anesthetic effects of barbiturates in this animal model. We postulate that the increase in the sensitivity to anesthetics results from a global suppression of excitatory neurotransmission in GluR2-deficient mice.

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Role of components of the phagocytic NADPH oxidase in oxygen sensing

Journal of Applied Physiology ◽

10.1152/japplphysiol.00564.2001 ◽

2002 ◽

Vol 93 (4) ◽

pp. 1357-1364 ◽

Cited By ~ 32

Author(s):

K. A. Sanders ◽

K. M. Sundar ◽

L. He ◽

B. Dinger ◽

S. Fidone ◽

...

Keyword(s):

Nadph Oxidase ◽

Carotid Body ◽

Minute Ventilation ◽

Mutant Mice ◽

Whole Body ◽

Type I ◽

Null Mutant ◽

Wild Type ◽

Null Mutant Mice

It has been hypothesized that O2sensing in type I cells of the carotid body and erythropoietin (EPO)-producing cells of the kidney involves protein components identical to the NADPH oxidase system responsible for the respiratory burst of phagocytes. In the present study, we evaluated O2sensing in mice with null mutant genotypes for two components of the phagocytic oxidase. Whole body plethysmography was used to study unanesthetized, unrestrained mice. When exposed to an acute hypoxic stimulus, gp91phox-null mutant and wild-type mice increased their minute ventilation by similar amounts. In contrast, p47phox-null mutant mice demonstrated increases in minute ventilation in response to hypoxia that exceeded that of their wild-type counterparts: 98.0 ± 18.0 vs. 20.0 ± 13.0% ( n = 11, P = 0.003). In vitro recordings of carotid sinus nerve (CSN) activity demonstrated that resting (basal) neural activity was marginally elevated in p47phox-null mutant mice. With hypoxic challenge, mean CSN discharge was 1.5-fold greater in p47phox-null mutant than in wild-type mice: 109.61 ± 13.29 vs. 72.54 ± 7.65 impulses/s ( n = 8 and 7, respectively, P = 0.026). Consequently, the hypoxia-evoked CSN discharge (stimulus-basal) was ∼58% larger in p47phox-null mutant mice. Quantities of EPO mRNA in kidney were similar in gp91phox- and p47phox-null mutant mice and their respective wild-type controls exposed to hypobaric hypoxia for 72 h. These findings confirm the previous observation that absence of the gp91phoxcomponent of the phagocytic NADPH oxidase does not alter the O2-sensing mechanism of the carotid body. However, absence of the p47phoxcomponent significantly potentiates ventilatory and chemoreceptor responses to hypoxia. O2sensing in EPO-producing cells of the kidney appears to be independent of the gp91phoxand p47phoxcomponents of the phagocytic NADPH oxidase.

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