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Biomedical subjects

M Philip

Publications and source records attributed to M Philip.

At least 19 recordsLinked to original sources
PubMed1992

Mutational analysis of third cytoplasmic loop domains in G-protein coupling of the HM1 muscarinic receptor.

We measured dose-response curves for carbachol stimulation of phosphatidyl inositol (PI) turnover with mutants of the Hm1 muscarinic cholinergic receptor having various deletions from amino acids 219 to 358 of the large third intracellular (i3) loop (208 to 366). These deletions had only small or no effects on the ability of Hm1 transfected into HEK 293 cells to stimulate PI turnover. This result indicates that only small regions of 9 to 11 amino acids adjacent to trans-membrane domains (TMDs) 5 and 6 can be directly involved in G protein coupling. Point mutations were constructed to test the role of charged amino acids in these junctions. A triple point mutation of Hm1 (E214 A/ E216K/ E221 K), which mimics the charge distribution in Hm2 (negatively coupled to cAMP) over the first 14 amino acids of i3, and a double point mutation in the N terminal junction, K359A/K361A, both failed to affect carbachol stimulated PI turnover. Therefore, charge distribution in the loop junctions appears to play a minor role in G protein coupling of Hm1 in HEK 293 cells.

Amino Acid Sequence↗
PubMed1992

Hm1 muscarinic cholinergic receptor internalization requires a domain in the third cytoplasmic loop.

Selected regions of the Hm1 muscarinic cholinergic receptor were mutated to analyze the molecular mechanisms of agonist-induced receptor internalization (or sequestration). The wild-type and mutant Hm1 genes were expressed, using pSG5, in U293 human kidney cells. Whereas surface receptor density measured with the polar tracer N-[3H]methylscopolamine was rapidly reduced by carbachol exposure, total receptor content measured with [3H]quinuclidinyl benzilate did not decline for at least 24 h, indicating the absence of extensive receptor down-regulation in U293 cells. Carbachol stimulation of phosphatidylinositol turnover paralleled receptor internalization, both with EC50 values of 10-20 microM. Furthermore, a D71N point mutation that prevented receptor activation also abolished carbachol-induced receptor internalization, indicating that receptor activation (but not necessarily second messenger stimulation) was required for internalization. Truncation of the COOH-terminal tail (K447 trunc) and point mutations of several potential Ser and Thr phosphorylation sites to Ala failed to affect receptor activation and internalization. In contrast, partial deletions of the third intracellular loop (i3) (Tyr208-Thr366) resulted in receptor mutants deficient in agonist-induced receptor internalization/sequestration. Various deletions caused either complete loss of internalization (d 232-358) or impaired internalization, ranging from 10 to 30% over 2 h, whereas wild-type Hm1 internalized to approximately 50%. Whereas the reason for the observed differences among the deficient deletion mutants remains unclear, the initial rate of N-[3H]methylscopolamine binding loss from the cell surface was much slower than that of wild-type Hm1 in each case. The deletion of only one single domain, 284-292 (SMESLTSSE), in the middle of i3 was consistently associated with impaired internalization. Domain 284-292 is partially conserved among closely related muscarinic receptors, whereas most of the remainder of i3 is not (except for the i3 membrane junctions), and similar Ser- and Thr-rich regions are present in many other G protein-coupled receptors. We propose that a small receptor domain in the middle of the i3 loop of Hm1 is involved in agonist-induced receptor internalization.

Amino Acid Sequence↗
PubMed1992

Mapping the ligand binding pocket of the human muscarinic cholinergic receptor Hm1: contribution of tyrosine-82.

The ligand binding pocket of many G protein-coupled receptors is thought to be located within the core formed by their seven transmembrane domains (TMDs). Previous results suggested that muscarinic antagonists bind to a pocket located toward the extracellular region of the TMDs, primarily at TMDs 2, 3, 6, and 7. Tyrosine-82 (Y82) is located in TMD2 only one helical turn from the presumed membrane surface of Hm1, whereas a phenylalanine (F124) is found in the equivalent position of the closely related Hm3. In order to determine the contribution of Y82 to Hm1 ligand binding and selectivity versus Hm3, we constructed the point mutation Y82 F of Hm1 and measured binding affinities of various ligands, with 3H-N-methylscopolamine (3H-NMS) as the tracer. The Hm1 wild-type receptor and the Y82F mutant were transfected into human embryonic kidney U293 cells. Whereas the affinities of NMS, carbachol, and atropine were either unchanged (carbachol) or enhanced by less than twofold (atropine and NMS), the affinity of the Hm1-selective pirenzepine was reduced threefold by the Y82F mutation. These changes parallel affinity differences of Hm1 and Hm3, indicating that the Y82 F mutation affects the binding pocket and that Y82 contributes to the binding selectivity among closely related muscarinic receptors.

Amino Acid Sequence↗
PubMed1992

Intraoperative monitoring of an unusual brachial plexus tumor.

This case illustrates the importance of intraoperative monitoring of neuronal function to help separate tumor tissue from neural tissue in a 54-year-old patient with left shoulder pain resulting from a desmoid tumor. Preoperative nerve conduction and electromyographic studies showed a lesion in the lateral cord of the brachial plexus, which was found to be intimately involved with the tumor mass and was splayed into a very thin effaced sheet of neural tissue. Stimulation of the tumor/nerve tissue mass proximal to the lesion was impossible due to the invasion of the brachial plexus by the tumor. The technique that was adapted for this unusual presentation was to stimulate the tumor/nerve tissue mass itself and record compound muscle action potentials distally. With the technique described, a subtotal resection of an aggressive fibromatosis enmeshed in the proximal brachial plexus was possible, and excellent relief of pain symptoms and retention of functional capabilities of the involved extremity were achieved.

Action Potentials↗
PubMed1990

Internalization of the Hm1 muscarinic cholinergic receptor involves the third cytoplasmic loop.

The m1 muscarinic receptor was previously shown to stimulate phosphatidyl inositol (PI) turnover and to internalize rapidly upon agonist activation. Three receptor mutants with large deletions of the third cytoplasmic loop (i3) of human Hm1, leaving only 11 and 8 amino acids at the amino and carboxy terminal junctions of i3, respectively, retained full ability to stimulate PI turnover, when expressed in U293 cells, but receptor internalization was greatly reduced in two mutants with deletions reaching close to the NH2 terminal of i3. We propose that a receptor domain located toward the amino terminal junction of i3 plays a role in Hm1 internalization.

Carbachol↗
PubMed1990

Structure and function of G protein coupled receptors.

The G protein coupled receptors (GPC-Rs) comprise a large superfamily of genes encoding numerous receptors which all show common structural features, e.g., seven putative membrane spanning domains. Their biological functions are extremely diverse, ranging from vision and olfaction to neuronal and endocrine signaling. The GPC-Rs couple via multiple G proteins to a growing number of recognized second messenger pathway, e.g., cAMP and phosphatidyl inositol turnover. This review summarizes our current knowledge of the molecular mechanisms by which the GPC-Rs activate second messenger systems, and it addresses their regulation and structure.

Amino Acid Sequence↗
PubMed1990

Reflex sympathetic dystrophy in central cord syndrome: case report and review of the literature.

Reflex sympathetic dystrophy (RSD) has been reported in incomplete spinal cord injury patients, most often occurring unilaterally; however to our knowledge, bilateral RSD has not been reported in patients with a central cord syndrome. We report a case of bilateral RSD in a patient with incomplete cervical myelopathy and the clinical picture of central cord syndrome. Diagnosis of RSD was based upon clinical, roentgenographic and scintigraphic findings. Management of RSD included elevation of forearm and hands, gentle active and passive range of movements of all upper extremity joints and systemic corticosteroids. With treatment, pain subsided, the range of motion of the joints improved and the patient achieved good functional recovery.

Aged↗
PubMed1990

Cardiac pacemaker inhibition by transcutaneous electrical nerve stimulation.

The use of transcutaneous electrical nerve stimulation (TENS) has increased in recent years for the management of chronic pain. One of its few contraindications is the presence of a cardiac pacemaker. Clinicians have recommended that cardiac monitoring be performed for a short interrupted period during the first application of TENS to ensure safety. We present two patients who were given trials of TENS for different chronic pain complaints. In both cases, electrocardiograms during the TENS trial did not reveal cardiac pacemaker dysfunction, but extended cardiac monitoring with the Holter monitor showed interference with pacemaker function. The abnormalities did not recur after reprogramming the sensitivity of the pacemakers. We suggest the use of extended cardiac monitoring for patients with cardiac pacemakers during the prolonged use of TENS to ensure safety and to determine any need for reprogramming of the pacemakers. Patients with cardiac pacemakers should not be excluded from the use of TENS, but careful evaluation and extended cardiac monitoring should be performed.

Aged↗
PubMed1989

Double disability of hemiplegia and hip disarticulation: rehabilitation outcome.

Double disability of hemiplegia and hip disarticulation is uncommon. To our knowledge, there are no reports of such patients and their rehabilitation outcomes. We report on a patient with hemiplegia and hip disarticulation who became independent in prosthetic ambulation and activities of daily living. Motivation, age, good neuromuscular status, and past successful prosthetic use favored the excellent rehabilitation outcome in this patient.

Adolescent↗
PubMed1985

Analysis of visual scanning patterns of pistol shooters.

Visual behaviour of pistol shooters of various levels of experience was studied using a video-oculometric technique to record the direction of gaze. A spatio-temporal analysis of the fixation patterns of subjects (five international pistol shooters and five international pentathletes) found that patterns were related to the level of skill. The pentathletes, whose level was lower, positioned their gaze on the weapon sight and visually followed the movement of the weapon towards the target. The pistol shooters appeared to use negative feedback in bringing up their weapons, so reducing the angle formed by the weapon, target and gaze. They fixed their gaze directly on the target or between the target and the weapon. Observations showed a prolonged time of attack (means = 1.107 s vs 0.558 s; P less than 0.001) and a shorter stabilization time (means = 0.714 s vs 1.216 s; P less than 0.001) for the most expert shooters. The psycho-physiological mechanisms which were the basis of these patterns and the reasons why shooters chose them are discussed.

Humans↗
PubMed1979

Nucleosome core histone complex isolated gently and rapidly in 2 M NaCl is octameric.

A relatively stable specific complex of the chromatin core histones H2A, H2B, H3, and H4 has been obtained in 2 M NaCl/25 mM sodium phosphate buffer, pH 7.0. The histone core complex has an apparent specific volume of 0.73 ml/g. Its sedimentation coefficient was dependent on rotor speed (angular velocity, omega) and attained different stable values at low and high rotor speeds. The drop in sedimentation coefficient occurred sharply between omega 2 values of about 9 x 10(6) and 1.1 x 10(7) (radians/sec)2. The s020,w corresponding to zero angular velocity (1 atmosphere pressure) was 6.6 S +/- (SEM) 0.1 S. At high rotor speeds the value decreased to 3.8 S +/- 0.1 S. The core complex has a diffusion coefficient, D20,w, of 5.4 x 10(-7) cm2/sec and a molecular weight of 108,000 +/- (SD) 2500.

Animals↗