Mechanisms of allosteric modulation at GABAB receptors by CGP7930 and GS39783: effects on affinities and efficacies of orthosteric ligands with distinct intrinsic properties

We determined the effects of the allosteric gamma-aminobutyric acid B receptor modulators CGP7930 and GS39783 on binding and function of orthosteric ligands with distinct intrinsic properties. In radioligand binding (saturation or displacement) experiments, the affinities of a number of competitive antagonists were decreased by the modulators, with no change in receptor number. The binding curves of the partial agonist CGP47656 comprised a high and a low affinity component; the affinity of the former was increased by the allosteric agents. The maximal stimulation of GTP[gamma](35)S binding via recombinant GABA(B) receptors by CGP47656 was increased 4-fold in the presence of 30 microM CGP7930 or GS39783. Two compounds known so far as "silent" competitive GABA(B) receptor antagonists, CGP35348 and 2-OH-saclofen, did not stimulate GTP[gamma](35)S binding on their own, but became low efficacy partial agonists in the presence of the two modulators. The potency of GABA to inhibit the formation of cAMP induced by a forskolin analog in a recombinant CHO cell line expressing GABA(B) receptors was increased by the modulators. CGP35348 and 2-OH-saclofen, like CGP47656, were partial agonists on their own in this assay, and the allosteric modulators increased the potency as well as the efficacy of all three compounds. With CGP52432, there was a trend towards inverse agonism in the cAMP assay. These results show that the intrinsic properties of orthosteric ligands are highly dependent on the characteristics of the assay system used and that allosteric modulators are useful tools for elucidating these properties.     

Partial brain dopamine D2 receptor agonists in the treatment of schizophrenia

Partial dopamine agonists showing high affinity but low efficacy at D2 receptors can act as dopaminergic "buffers," reducing dopaminergic activity when it is excessive, and promoting it when reduced. This makes them of interest as potential therapeutic agents for the treatment of both positive and negative symptoms in schizophrenia, where they should also result in fewer or less severe motor disturbances than classical neuroleptics. SDZ 208-911 and SDZ 208-912 are amino-ergolines exhibiting partial agonistic properties in the rat, where they inhibit apomorphine-induced stereotypies, are only weakly cataleptogenic, induce varying degrees of circling behavior after unilateral lesioning of the nigrostriatal pathway, and strongly suppress prolactin secretion. The least agonistically acting agent, SDZ 208-912, should be effective against positive symptoms, whereas SDZ 208-911 could be suitable for the treatment of negative symptoms. In addition to possible therapeutic effects, the clinical testing of this class of agent should help to elucidate the status of central dopaminergic function in schizophrenic psychosis.     

Evaluation of a novel needle guide for ultrasound-guided phantom vessel cannulation

We evaluated a novel, sled-mounted needle guide for ultrasound-guided vessel cannulation. Fifty medical students were randomly assigned to use ultrasound with the sled (sled group, n = 23) or ultrasound without the sled (control group, n = 27) for vessel cannulation in a phantom. For each of 15 attempts we recorded cannulation time and designated a successful cannulation as 1 and a failure as 0. Our primary outcome was the mean overall success rate. The median (IQR [range]) number of successes in the sled group and control group were 15.0 (13.0-15.0 [11.0-15.0]) and 11.0 (9.0-13.0 [6.0-15.0]), respectively (p < 0.001). Cannulation time decreased from the first to the last attempt in the sled group from 7.0 s (6.0-10.0 [4.0-16]) s to 4.0 s (3.0-4.0 [1.0-6.0]) s and in the control group from 35.0 s (27.0-35.0 [11.0-35.0]) s to 7.0 s (5.0-10.0 [3.0-25.0]) s. The sled group demonstrated a shorter cannulation time at each attempt (p < 0.001). The novel sled improved the success rate and efficiency of ultrasound-guided phantom vessel cannulation.     

Roles of GABA<Subscript>B</Subscript> receptor subtypes in presynaptic auto- and heteroreceptor function regulating GABA and glutamate release

Γ-Aminobutyric acid B (GABA_B) receptors are heterodimers composed of two subunits GABA_B(1) and GABA_B(2), the former existing in two isoforms GABA_B(1a) and GABA_B(1b). The contributions of individual receptor subunits and isoforms to GABA_B auto- and heteroreceptor functions were investigated, using release experiments in cortical slice preparations from corresponding knockout mice. Presynaptic GABA_B autoreceptors are located on GABAergic terminals and inhibit GABA release, whereas presynaptic GABA_B heteroreceptors control the release of other neurotransmitters (e.g. glutamate). Neither baclofen nor the selective antagonist CGP55845 at maximally active concentrations affected [³H]GABA release in slices from GABA_B(1)−/− mice. The amount of [³H]GABA released per pulse was unaffected by the stimulation frequency in slices from GABA_B(1)−/− and GABA_B(2)−/− demonstrating a loss of GABA_B autoreceptor function in these knockout animals. The GABA_B receptor agonist baclofen was ineffective in modulating glutamate release in cortical slices from GABA_B(2)−/− mice, showing that heteroreceptor function was abolished as well. Next we investigated knockout mice for the two predominant GABA_B(1) isoforms expressed in brain, GABA_B(1a) and GABA_B(1b). In cortical, hippocampal and striatal slices from both GABA_B(1a)−/− and GABA_B(1b)−/− mice, the frequency dependence of [³H]GABA released per pulse was maintained, suggesting that both isoforms participate or can substitute for each other in GABA_B autoreceptor function. By contrast, the efficacy of baclofen to inhibit glutamate release was substantially reduced in GABA_B(1a)−/−, but essentially unaltered in GABA_B(1b)−/− mice. Our data suggest that functional GABA_B heteroreceptors regulating glutamate release are predominantly, but not exclusively composed of GABA_B(1a) and GABA_B(2) subunits. An erratum to this article can be found at     

Genotyping the Butyrylcholinesterase in Patients with Prolonged Neuromuscular Block after Succinylcholine

Background: Succinylcholine remains the standard neuromuscular blocking drug for tracheal intubation in emergency situations. The short duration of action is due to its rapid hydrolytic degradation by butyrylcholinesterase (plasmacholinesterase). Multiple variants of this enzyme are known (A, F, S, H, J, K variants) with different effects on enzyme activity. This study was undertaken to evaluate the use of molecular genetic methods in patients with clinically prolonged neuromuscular block.

Methods: Nine patients with a neuromuscular block of 14 min to 5 h were selected. All four exons of the butyrylcholinesterase were amplified by polymerase chain reaction and analyzed by automated sequencing. Molecular genetic results were compared with clinical relaxation time and with biochemical test results (total butyrylcholinesterase activity, dibucaine and fluoride inhibition).

Results: Seven of nine patients were mutation carriers. Five of these had more than one mutation. The A and K variants were the most frequent variations. Three of four patients who were homozygous for the A variant were also carriers of the K allele. The authors identified one novel mutation (G1294T) introducing a stop codon at amino acid position 432. The duration of neuromuscular block was substantially different between patients with identical butyrylcholinesterase genotypes.     

In Vivo Whole-Body Resting Energy Expenditure and Insulin Action in Human Malignant Hyperthermia

Background: Malignant hyperthermia (MH) is a pharmacogenetic disease triggered by volatile anesthetics or succinylcholine. The disorder is heterogenetic and caused by abnormal calcium regulation within skeletal muscle cells. No clear metabolic differences have been found in MH-susceptible (MHS) persons in vivo while not having MH episodes, but some reported signs suggest that insulin action and energy turnover might be altered in muscle of MHS persons.

Methods: In fasting and insulin-stimulated conditions, using the glucose clamp technique and indirect calorimetry, we assessed in vivo resting energy expenditure (REE) and nutrient utilization rates in 10 MHS, 5 MH-equivocal (MHE) and 10 MH-negative (MHN) persons from 14 families. With a model using the persons' fat-free mass, fat mass, age, and gender, we calculated their predicted REE and compared it with measured REE in 10 MHS and 10 MHN persons (measured - predicted = residual REE).

Results: In vivo measured REE and glucose disposal rates were similar in 10 MHS and 10 MHN persons. Only during insulin stimulation was residual REE greater in MHS persons (6.4%;P = 0.013).     

JP‐45/JSRP1 Variants Affect Skeletal Muscle Excitation–Contraction Coupling by Decreasing the Sensitivity of the Dihydropyridine Receptor

JP‐45 (also JP45; encoded by JSRP1) is an integral protein constituent of the skeletal muscle sarcoplasmic reticulum junctional face membrane interacting with Ca_v1.1 (the α.1 subunit of the voltage‐sensing dihydropyridine receptor, DHPR) and the luminal calcium‐binding protein calsequestrin. Two JSRP1 variants have been found in the human population: c.323C>T (p.P108L) in exon 5 and c.449G>C (p.G150A) in exon 6, but nothing is known concerning the incidence of these polymorphisms in the general population or in patients with neuromuscular diseases nor the impact of the polymorphisms on excitation–contraction (EC) coupling. In the present report, we investigated the frequencies of these two JSRP1 polymorphisms in the Swiss malignant hyperthermia population and studied the functional impact of the variants on EC coupling. Our results show that the polymorphisms are equally distributed among malignant hyperthermia negative, malignant hyperthermia equivocal, and malignant hyperthermia susceptible individuals. Interestingly, however, the presence of either one of these JP‐45 variants decreased the sensitivity of the DHPR to activation. The presence of a JSRP1 variant may explain the variable phenotype seen in patients with malignant hyperthermia carrying the same mutation and, more importantly, may counteract the hypersensitivity of EC coupling caused by mutations in the RYR1 gene.