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Biomedical subjects

C Hammond

Publications and source records attributed to C Hammond.

At least 37 records · Page 2Linked to original sources
PubMed1998

Differential management of Ca2+ oscillations by anterior pituitary cells: a comparative overview.

Most electrical and ionic properties of anterior pituitary cells are common to all pituitary cell types; only gonadotropes exhibit a few cell specific features. Under basal conditions, the majority of pituitary cells in vitro, irrespective of their cell type, display spontaneous action potentials and [Ca2+]i transients that result from rhythmic Ca2+ entry through L-type Ca2+ channels. The main function of these action potentials is to maintain cells in a readily activable responsive state. We propose to call this state a 'pacemaker mode', since it persists in the absence of extrinsic stimulation. When challenged by hypothalamic releasing hormones, cells exhibit two distinct response patterns: amplification of pacemaker activity or shift to internal Ca2+ release mode. In the internal Ca2+ release mode, [Ca2+]i oscillations are not initiated by entry of external Ca2+, but by release of Ca2+ from intracellular stores. In somatotropes and corticotropes, GHRH or CRH triggers the pacemaker mode in silent cells and amplifies it in spontaneously active cells. In contrast, in gonadotropes GnRH activates the internal Ca2+ release mode in silent cells and switches already active cells from the pacemaker to the internal Ca2+ release mode. Interestingly, homologous normal and tumoral cells display the same type of activity in vitro, in the absence or presence of hypothalamic hormones. Pacemaker and internal Ca2+ release modes are likely to serve different purposes. Pacemaker activity allows long-lasting sequences of [Ca2+]i oscillations (and thus sustained periods of secretion) that stop under the influence of hypothalamic inhibitory peptides. In contrast, the time during which cells can maintain internal Ca2+ release mode depends upon the importance of intracellular Ca2+ stores. This mode is thus more adapted to trigger secretory peaks of large amplitude and short duration. On the basis of these observations, theoretical models of pituitary cell activity can be proposed.

Action Potentials↗
PubMed1998

Referrals for depression by primary care physicians. A pilot study.

BACKGROUND: Communication between primary care physicians (PCPs) and mental health providers (MHPs) is integral to the management of depressive disorders. Our study investigated referrals from PCPs to MHPs in a rural research network. METHODS: From April 1997 to September 1997, 5 family physicians tracked 6 weeks of referrals for depression and non-mental health problems and assessed the outcomes of these referrals after 3 months. The referrals were characterized by the nature and extent of communication between the PCP and the MHP and by the effectiveness of the consultation. RESULTS: Sixty-seven patients (44 with non-mental health disorders and 23 with depressive disorders) were identified and followed. Analysis of the initial referral process showed that the referring physicians felt a greater sense of urgency for the referrals for depression. Written evidence of the referral in the patient's chart at the 3-month survey was more common for non-mental health disorders. CONCLUSIONS: This pilot study demonstrates that there are communication barriers between PCPs and their mental health colleagues. Enhanced communication might improve satisfaction with the referral process. The details of the referral process need further study.

Communication↗
PubMed1997

Negative regulation by HLA-DO of MHC class II-restricted antigen processing.

HLA-DM is a major histocompatibility complex (MHC) class II-like molecule that facilitates antigen processing by catalyzing the exchange of invariant chain-derived peptides (CLIP) from class II molecules for antigenic peptides. HLA-DO is a second class II-like molecule that physically associates with HLA-DM in B cells. HLA-DO was shown to block HLA-DM function. Purified HLA-DM-DO complexes could not promote peptide exchange in vitro. Expression of HLA-DO in a class II+ and DM+, DO- human T cell line caused the accumulation of class II-CLIP complexes, indicating that HLA-DO blocked DM function in vivo and suggesting that HLA-DO is an important modulator of class II-restricted antigen processing.

Amino Acid Sequence↗
PubMed1997

Growth hormone-releasing hormone triggers pacemaker activity and persistent Ca2+ oscillations in rat somatotrophs.

1. The effects of brief applications of growth hormone-releasing hormone (GHRH) to male rat somatotrophs in culture were analysed with the perforated patch clamp technique to record changes in potential or with fura-2 imaging techniques to measure variations of cytosolic Ca2+ concentration ([Ca2+]i). 2. Silent somatotrophs (n = 61) had a mean resting potential of -37 +/- 1 mV and a mean basal [Ca2+]i of 30 +/- 4 nM. Brief GHRH applications (30 nM, 40 s) triggered rhythmic action potentials (23.6 +/- 0.9 mV, 613 +/- 82 ms, 0.21 +/- 0.02 Hz) and [Ca2+]i increase (to 352 +/- 30 nM) followed by rhythmic [Ca2+]i transients (to 138 +/- 6 nM) that persisted up to 90 min after the last GHRH application. Both action potentials and [Ca2+]i transients were totally and reversibly blocked by removing external Ca2+ or Na+ or by adding inorganic Ca2+ channel blockers or nifedipine (3 microM). 3. Somatostatin (1-300 nM), carbamylcholine (0.1-1 microM) and muscarine (0.1-1 microM) each had a dose-dependent inhibitory effect, from a decrease of Ca2+ spike duration and frequency to a complete block of the GHRH-evoked action potentials. 4. The present results show that somatotrophs in culture have intrinsic membrane properties that allow them to sustain a pacemaker activity and subsequent long-lasting sequences of [Ca2+]i oscillations triggered by short pulses of GHRH and inhibited by somatostatin and muscarinic agonists.

Action Potentials↗
PubMed1997

Misfolded major histocompatibility complex class I heavy chains are translocated into the cytoplasm and degraded by the proteasome.

N-acetyl-L-leucyl-L-leucyl-L-norleucinal (LLnL), which reversibly inhibits the proteasome in addition to other proteases, and a more specific irreversible inhibitor of the proteasome, lactacystin, were found to cause the accumulation of major histocompatibility complex (MHC) class I heavy chains in the cytosol of the beta2-microglobulin-deficient cell line Daudi and the TAP-deficient cell line .174. These cell lines, which are severely impaired in their ability to fold MHC class I heavy chain, showed an accumulation of soluble class I heavy chains at different rates over a period of hours in the presence of LLnL. The accumulation of soluble class I heavy chains in the presence of either LLnL or lactacystin was easily revealed in Daudi and .174 but almost undetectable in a Daudi transfectant expressing beta2-microglobulin and in 45.1, the wild-type parent of .174. The soluble class I heavy chain was also found to be devoid of its N-linked glycan and to be located in the cytosol. When the gene for ICP47, a herpes simplex virus protein that blocks the translocation of peptides into the endoplasmic reticulum, was transfected into 45.1, a similar accumulation of soluble MHC class I heavy chain was detectable. These data suggest that in cells where the MHC class I molecule is unable to assemble properly, the misfolded heavy chain is removed from the endoplasmic reticulum to the cytosol, deglycosylated, and degraded by the proteasome.

Acetylcysteine↗
PubMed1996

HLA-DM interactions with intermediates in HLA-DR maturation and a role for HLA-DM in stabilizing empty HLA-DR molecules.

Major histocompatibility complex (MHC) class II-positive cell lines which lack HLA-DM expression accumulate class II molecules associated with residual invariant (I) chain fragments (class II-associated invariant chain peptides [CLIP]). In vitro, HLA-DM catalyzes CLIP dissociation from class II-CLIP complexes, promoting binding of antigenic peptides. Here the physical interaction of HLA-DM with HLA-DR molecules was investigated. HLA-DM complexes with class II molecules were detectable transiently in cells, peaking at the time when the class II molecules entered the MHC class II compartment. HLA-DR alpha beta dimers newly released from I chain, and those associated with I chain fragments, were found to associate with HLA-DM in vivo. Mature, peptide-loaded DR molecules also associated at a low level. These same species, but not DR-I chain complexes, were also shown to bind to purified HLA-DM molecules in vitro. HLA-DM interaction was quantitatively superior with DR molecules isolated in association with CLIP. DM-DR complexes generated by incubating HLA-DM with purified DR alpha beta CLIP contained virtually no associated CLIP, suggesting that this superior interaction reflects a prolonged HLA-DM association with empty class II dimers after CLIP dissociation. Incubation of peptide-free alpha beta dimers in the presence of HLA-DM was found to prolong their ability to bind subsequently added antigenic peptides. Stabilization of empty class II molecules may be an important property of HLA-DM in facilitating antigen processing.

Amino Acid Sequence↗
PubMed1996

Trafficking of major histocompatibility complex class II molecules through intracellular compartments containing HLA-DM.

The endosomal site(s) where MHC class II molecules become competent to bind antigenic peptide has not been completely characterized. We identified endocytic compartments through which newly synthesized MHC class II molecules move prior to their expression on the plasma membrane. The compartments co-sediment with lysosomes in the most dense regions of Percoll gradients. The appearance of proteolytic fragments of the invariant chain (I chain), namely leupeptin-induced proteins (LIPs) and class-II-associated invariant chain peptides (CLIP), in this region of the gradient suggests that the release of MHC class II molecules from I chain association occurs within these vesicles. The formation of SDS-stable alpha beta dimers indicated that MHC class II molecules contained within these compartments are receptive to peptide binding. A majority of the HLA-DM protein was found in the same region of the Percoll gradient, consistent with its established function in MHC class-II-restricted antigen presentation. Immunoelectron micrographs of dense-sedimenting compartments indicated that I chain, MHC class II, and DM molecules are contained within both multivesicular and multilamellar vesicles. The final stages of I chain dissociation from MHC class II molecules and DM-mediated peptide loading probably occur in these compartments.

Antigen Presentation↗
PubMed1996

HLA-DM is localized to conventional and unconventional MHC class II-containing endocytic compartments.

HLA-DM molecules remove invariant (Ii) chain peptides from newly synthesized MHC class II complexes. Their localization may thus delineate compartments, e.g., MIIC, specialized for loading peptides onto class II molecules. In murine A20 B cells, however, DM is not restricted to specialized endosomal class II-containing vesicles (CIIV). Although DM was found in CIIV, it was also found throughout the endocytic pathway, principally in lysosomes devoid of class II molecules. In human lymphoblasts, HLA-DM was found in structures indistinguishable from late endosomes or lysosomes, although in these cells the lysosomes contained MHC class II molecules. Thus, the distribution of HLA-DM does not necessarily identify specialized class II compartments. Many "MIIC" may represent conventional lysosomes that accumulate MHC class II and HLA-DM in a number of cell types.

Animals↗
PubMed1995

Folding and oligomerization of influenza hemagglutinin in the ER and the intermediate compartment.

Influenza hemagglutinin (HA) was used to analyze the stepwise folding and oligomeric assembly of glycoproteins in the early secretory pathway of living cells. In addition to mature trimers, six distinct maturation intermediates were identified. Of these, all the incompletely oxidized forms were located in the endoplasmic reticulum (ER) and associated with calnexin, a membrane-bound, lectin-like ER chaperone. Once fully oxidized, the HA dissociated from calnexin as a monomer, which rapidly became resistant to dithiothreitol (DTT) reduction. Part of these extensively folded molecules moved as monomers into the intermediate compartment between the ER and the Golgi complex. Assembly of homotrimers occurred without calnexin-involvement within the ER and in the intermediate compartment. When anchor-free HA molecules were analyzed, it was found that they reach the DTT-resistant monomeric conformation but fail to trimerize. Taken together, the results provide a definition and intracellular localization of several intermediates in the conformational maturation of HA, including the immediate precursor for trimer assembly.

Animals↗
PubMed1995

Quality control in the secretory pathway.

Whereas newly synthesized proteins that have acquired a properly folded and assembled structure are transported from the endoplasmic reticulum to their final destinations, incompletely folded and assembled proteins are, as a rule, retained and eventually degraded. The molecular mechanisms of this unique molecular sorting phenomenon, called 'quality control', have been illuminated by recent studies.

Animals↗
PubMed1995

The effect of head and neck irradiation on taste dysfunction: a prospective study.

Taste loss is a major cause of morbidity in patients undergoing head and neck irradiation. In a prospective study, 26 patients undergoing radical head and neck irradiation at the Royal Marsden Hospital, Sutton, and the Queen Elizabeth Hospital, Birmingham, were assessed for taste loss and xerostomia. Taste was tested using a subjective questionnaire and by objective taste testing with a series of solute solutions (sucrose, sodium chloride, urea and hydrochloric acid) at increasing concentrations, to determine the threshold level of taste sensation, both before and after radiotherapy. Xerostomia was assessed using a patient questionnaire. The volume of tongue and parotid contained within the high dose volume of the radiation treatment field was determined for each patient and correlated with the degree of objective and subjective taste loss as well as the degree of xerostomia. The results have shown that both objective (r = 0.59; P = 0.0016) and subjective taste loss (r = 0.78; P = 0.0001) was significantly associated with the proportion of tongue, but not parotid, contained within the radiation treatment field. The data gave no evidence to suggest any relationship between recovery of taste loss and volume of parotid or tongue irradiated. However, recovery of subjective taste loss, 1 month after completing radiotherapy was seen in two patients, both of whom had been treated using a wedge pair technique to avoid the contralateral area of the tongue. Changes in xerostomia were significantly correlated with the proportions of both tongue (r = 0.54; P = 0.004) and parotid (r = 0.82; P = 0.0001) within the radiation treatment fields.(ABSTRACT TRUNCATED AT 250 WORDS)

Head and Neck Neoplasms↗
PubMed1994

Folding of VSV G protein: sequential interaction with BiP and calnexin.

The endoplasmic reticulum (ER) contains molecular chaperones that facilitate the folding of proteins in mammalian cells. Biosynthetic labeling was used to study the interactions of two chaperones, BiP and calnexin, with vesicular stomatitis virus (VSV) glycoprotein (G protein). Coimmunoprecipitation of G protein with the chaperones showed that BiP bound maximally to early folding intermediates of G protein, whereas calnexin bound after a short lag to more folded molecules. Castanospermine, an inhibitor of ER glucosidases, blocked the binding of proteins to calnexin and inhibited G protein folding. Interaction with calnexin was necessary for efficient folding of G protein and for retention of partially folded forms.

Animals↗
PubMed1994

Role of N-linked oligosaccharide recognition, glucose trimming, and calnexin in glycoprotein folding and quality control.

Using a pulse-chase approach combined with immunoprecipitation, we showed that newly synthesized influenza virus hemagglutinin (HA) and vesicular stomatitis virus G protein associate transiently during their folding with calnexin, a membrane-bound endoplasmic reticulum (ER) chaperone. Inhibitors of N-linked glycosylation (tunicamycin) and glucosidases I and II (castanospermine and 1-deoxynojirimycin) prevented the association, whereas inhibitors of ER alpha-mannosidases did not. Our results indicated that binding of these viral glycoproteins to calnexin correlated closely with the composition of their N-linked oligosaccharide side chains. Proteins with monoglucosylated oligosaccharides were the most likely binding species. On the basis of our data and existing information concerning the role of monoglucosylated oligosaccharides on glycoproteins, we propose that the ER contains a unique folding and quality control machinery in which calnexin acts as a chaperone that binds proteins with partially glucose-trimmed carbohydrate side chains. In this model glucosidases I and II serve as signal modifiers and UDP-glucose:glycoprotein glucosyltransferase, as a folding sensor.

Animals↗
PubMed1994

Anoxic LTP sheds light on the multiple facets of NMDA receptors.

Hippocampal neurones in the CA1 region have become a model system to study the mechanisms of long-term potentiation (LTP) and memory processes. The CA1 region is also highly vulnerable to ischaemic or anoxic episodes which induce a selective and delayed degeneration of pyramidal neurones. In CA1 neurones, anoxic episodes generate a novel form of LTP to which we refer as anoxic LTP. In common with tetanic LTP, the induction of anoxic LTP is voltage- and NMDA receptor-dependent. However, in contrast with tetanic LTP, the expression of anoxic LTP is mediated exclusively by NMDA receptors. These observations suggest that anoxic-ischaemic episodes trigger a switch in favour of NMDA receptor-operated synaptic transmission. We suggest that the multiple forms of NMDA receptor-dependent LTPs are determined by extracellular and intracellular modulatory sites of this receptor.

Animals↗
PubMed1994

Quality control in the secretory pathway: retention of a misfolded viral membrane glycoprotein involves cycling between the ER, intermediate compartment, and Golgi apparatus.

Proteins synthesized in the ER are generally transported to the Golgi complex and beyond only when they have reached a fully folded and assembled conformation. To analyze how the selective retention of misfolded proteins works, we monitored the long-term fate of a membrane glycoprotein with a temperature-dependent folding defect, the G protein of tsO45 vesicular stomatitis virus. We used indirect immunofluorescence, immunoelectron microscopy, and a novel Nycodenz gradient centrifugation procedure for separating the ER, the intermediate compartment, and the Golgi complex. We also employed the folding and recycling inhibitors dithiothreitol and AIF4-, and coimmunoprecipitation with calnexin antibodies. The results showed that the misfolded G protein is not retained in the ER alone; it can move to the intermediate compartment and to the cis-Golgi network but is then recycled back to the ER. In the ER it is associated with calnexin and BiP/GRP78. Of these two chaperones, only BiP/GRP78 seems to accompany it through the recycling circuit. Thus, the retention of this misfolded glycoprotein is the result of multiple mechanisms including calnexin binding in the ER and selective retrieval from the intermediate compartment and the cis-Golgi network.

Aluminum Compounds↗
PubMed1994

Glutamate metabotropic receptors increase a Ca(2+)-activated nonspecific cationic current in CA1 hippocampal neurons.

1. We studied the currents evoked in CA1 pyramidal neurons by the selective metabotropic glutamate receptor (mGluR) agonist 1S,3R-1-aminocyclopentane-1,3-dicarboxylate (1S,3R-ACPD; 100 microM, 2.30-5 min) with the single-electrode voltage-clamp technique in the continuous presence of tetrodotoxin (1 microM), bicuculline (10 microM), 6-cyano-7-nitroquinoxaline-2,3-dione (15 microM), and D-2-amino-5-phosphonovaleric acid (50 microM) to depress action potentials and synaptic activity. Microelectrodes were filled with 3M CsCl or 2 M Cs2SO4. 2. With CsCl-filled microelectrodes, bath application of 1S,3R-ACPD induced an inward current of -308 +/p 50 (SE) pA amplitude [holding potential (VH -60 mV, n = 12)] associated with a conductance decrease (26.5 +/- 5.6%, P < or = 0.0022, n = 12). The current-voltage (I-V) relation of the 1S,3R-ACPD-induced (difference) current investigated using ramp voltage commands from -130 to +10 mV had a V shape with two reversal potentials: -99.6 +/- 3.4 and -17.5 +/- 3.0 mV (n = 12). 3. In contrast, in the presence of external K+ channel blockers (2 mM Ba2+ and 6 mM Cs+ or 25 mM tetraethylammonium, 6 mM Cs+, and 3 mM 4-aminopyridine), 1S,3R-ACPD also generated an inward current, albeit of smaller amplitude (-114.2 +/- 27.5 pA, P < or = 0.003, VH -60 mV, n = 8). This current was associated with a conductance increase (20.7 +/- 3.1%, P < or = 0.0117, n = 8), decreased linearly with depolarization (from -130 to -60 mV), and reversed polarity at an estimated potential of -20.7 +/- 3.6 mV (n = 8). We refer to this current recorded in the presence of K+ channel blockers as IACPD. 4. In the presence of Cd2+ (200 microM, to block voltage-dependent Ca2+ channels that are readily activated in the presence of K+ channel blockers) and a low Ca2+ concentration (100 microM), IACPD decreased linearly from -130 to +10 mV and reversed polarity at -15.8 +/- 8.5 mV (n = 5).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗