Biomedical subjects

P Brodin

Publications and source records attributed to P Brodin.

At least 37 records · Page 2Linked to original sources

PubMed1995

Effects of triclosan on the rat phrenic nerve-diaphragm preparation.

It has been reported that the lipid soluble, anti-inflammatory drug triclosan, which is currently used in toothpastes and mouthrinses, may reduce pain. This may be an aspect of the anti-inflammatory effect of triclosan, which probably reduces the production of prostaglandin PGE2. However, triclosan may also exhibit a direct analgesic effect through an effect on excitable membranes. The aim of the present study was to examine the possible effect of triclosan on the nerve action potential, on the neuromuscular transmission and on the excitation-contraction coupling of skeletal muscle. Phrenic nerve-hemidiaphragm preparations, as well as isolated phrenic nerves, were dissected from Wistar albino rats and mounted by standard methods in Tyrode solution. Both during indirect and direct stimulation, the preparations were irreversibly inhibited by triclosan at concentrations higher than 5.0 x 10(-6)M. At low concentrations, the inhibition of the twitches during indirect stimulation and of the compound action potential was probably caused by a threshold increase for excitation of the phrenic nerve. At high concentrations, an additional inhibitory effect at the neuromuscular junction was disclosed. The directly stimulated preparation was also probably inhibited by an increase of the threshold for initiation of the muscle action potentials. In addition, a reduced KCl contracture and an acceleration of the caffeine contracture indicated an interaction with the sarcolemma. These results suggest that the analgesic effect of triclosan may be due to a direct interaction with excitable membranes.

Action Potentials↗

PubMed1995

The effects of low level laser treatment on recovery of nerve conduction and motor function after compression injury in the rat sciatic nerve.

An animal study is presented examining the effect of low level laser (LLL) treatment on nerve regeneration following axonotmesis. Twenty animals received a standardised injury to the right sciatic nerve using a time, load and length sequence (10 min, 150 N, 5 mm) known to cause extensive axonal degeneration of the rat sciatic nerve. The LLL treatment was administered using a hand-held laser probe in light contact with the skin on the dorsal aspect of the hind leg overlying the site of the axonotmesis injury to the sciatic nerve. A group of 10 animals were treated with 6J of LLL (GaAlAs 830 nm) daily for a period of 28 d. Ten more animals were treated daily with a sham exposure setting and served as controls. Nerve function was assessed by a recognised method of walking tract print analysis; the "Sciatic Functional Index" (SFI), and nerve regeneration was assessed by recording the evoked compound action potentials (cAP) in the common peroneal nerve. At 21 d post-injury, the laser-treated group had a significantly lower median SFI than the sham laser-treated group, indicating that the real laser treatment had improved functional recovery in the nerve. However, no differences were found between the evoked cAP parameters that were measured in the laser-treated and sham laser-treated groups. Histological examination reiterated the lack of difference between the two groups. Consequently, the effects of LLL on recovery must have occurred more peripherally to the point measured.

Animals↗

PubMed1995

Effects of mandibular nerve block on heat- or cold-induced changes in pulpal blood flow in man.

Laser Doppler flowmetry (LDF) was used to study the effect of mandibular nerve block, using 3% mepivacain, on heat- or cold-induced changes in pulpal blood flow (PBF) evoked by application of cold or heat to the lingual surfaces of teeth 33 and 43 in nine young subjects. PBF on average showed a 7% increase in response to heat (48 degrees C) and a 20% decrease when exposed to cold (3 degrees C). Neither response was affected by mandibular block. From this we concluded that the effects on PBF of the presently applied heating and cooling of the tooth were solely due to direct thermal influences on the blood vessels, without involvement of vasomotor or sensory nerves.

Adult↗

PubMed1994

Reflex responses of motor units in human masseter muscle to mechanical stimulation of a tooth.

The reflex responses evoked by controlled mechanical stimulation of an upper central incisor tooth in single motor units in the human masseter muscle were examined. The stimuli were (brisk) taps and (slow) pushes of about 2 N peak force, applied orthogonally to the labial surface of the ipsilateral upper central incisor tooth. The reflex responses of the motor units were characterised by analysis of the changes in the durations of the first and second interspike intervals (ISIs) immediately following the stimulus. A significant increase in the duration of these ISIs in comparison with pre-stimulus ISIs indicated inhibition, and significant shortening indicated excitation. Twenty masseter motor units were tested with both the pushes and the taps. The brisk taps elicited a significant reflex inhibition in 16 of the 20 motor units at a latency of 13 ms and duration of 37 ms. This inhibition was followed by significant excitation in 11 of the 20 units at latencies of 71 ms, lasting for 29 ms. The short-latency response to slow pushes was significant inhibition in four units: significant excitation in one unit and no response in 15 units. The slow pushes evoked a significant long-latency excitatory reflex response in 12 of the 20 units at latencies of 77 ms and lasting for 40 ms. The shapes and amplitudes of the compound post-synaptic potentials underlying the reflex responses in the motoneurones were estimated. It is concluded that stimulation of periodontal mechanoreceptors usually activates an excitatory reflex pathway to the jaw-closing motoneurones. This probably helps to grip the food bolus between the teeth during chewing. However, when the rate of application of the stimulus is large enough, a short-latency inhibitory response is evoked which, if of sufficient duration, may over-ride the subsequent excitatory response. Inhibition of the jaw-closing muscles will tend to protect the teeth and soft tissues when one bites unexpectedly on a hard object while chewing.

Adult↗

PubMed1994

Evocation of either excitatory or inhibitory reflex responses in human masseter muscle by electrical stimulation of the lip at varying intensities.

Electrical stimuli at 1 Hz with pulse widths of 0.05, 0.1 and 1 ms with intensities from two to six times sensory threshold (2-6 T) were delivered to the lower lip. The reflex responses were monitored by surface electromyography of the ipsilateral masseter muscle. An excitatory response that was not preceded or followed by an inhibition could be evoked in seven out of ten subjects at intensities below 5 T at all pulse widths. A higher stimulus intensities, the excitation disappeared and/or was preceded by as short-latency inhibition (SLI) or a long-latency inhibition (LLI). The electrical threshold for the excitatory response was statistically lower than the SLI and LLI, especially when longer pulse widths wee used. Three subjects demonstrated a primarily excitatory response, whereas four had a more pronounced inhibitory response. It was concluded that separate populations of myelinated fibres may be responsible for the responses: the lowest-threshold fibres may elicit excitatory responses and fibres with higher thresholds may evoke inhibitory responses. Another possible explanation is that central spatial summation could be responsible for the opening of the inhibitory and excitatory central pathways. The excitatory response may be the result of a reflex pathway similar to that evoked by activation of periodontal mechanoreceptors, and could be responsible for the load compensation mechanisms during chewing and/or positioning of food. The inhibitory responses are well known, and are considered to be a protective reflex.

Adolescent↗

PubMed1994

Effects of cooling and heating of the tooth on pulpal blood flow in man.

Laser Doppler flowmetry (LDF) was used to study the changes in pulpal blood flow (PBF) evoked by application of cold or heat to the palatinal surfaces of teeth 11 or 21 in nine young subjects. Switching from a thermode temperature of 33 degrees C to 5 degrees C on average induced a slow decrease of PBF to about 80% of control, and also warming to 39 degrees C evoked a small reduction in most subjects. Interindividual differences were large, however, and both cooling and warming sometimes triggered a rise in PBF. In contrast, skin blood flow, as recorded with LDF in the forearm, invariably rose during warming and fell during local cooling. The results suggested a more complex interaction between local and nervously mediated effects of moderate changes in temperature in the tooth pulp than skin, and that the previously held view of cold and heat decreasing and increasing PBF, respectively, is wrong.

Adult↗

PubMed1994

Physiological regulation of the secretion of histatins and statherins in human parotid saliva.

The small salivary phosphoproteins, histatins and statherins, have important functions in the oral cavity in terms of antimicrobial actions and regulation of calcium phosphate homeostasis. Neither the effects of various physiological stimuli on their secretion nor the nature of the efferent receptor involved in the stimulus-secretion coupling has been determined previously. These aspects are important for improved understanding of the secretory control of salivary proteins and may have implications regarding the effects of specific medications on salivary constituents and oral health. The effects of graded mechanical (chewing on short and long silicone tubings) and gustatory stimulation (0.5, 1.5, and 5.0% citric acid) on the secretion of histatins and statherins were studied in the presence and absence of adrenolytic agents (n = 10). In this model, secretory rates of both proteins increased with increases in flow rate, with 5.0% citric acid representing a particularly potent stimulus. Histatin and statherin secretory rates were significantly reduced by the beta 1-adrenolytic agent (histatins to 58 to 72% and statherins to 11 to 29% of that in corresponding control experiments), but not by the alpha 1-adrenolytic agent. Since the beta 1-adrenergic receptors played an important role in the stimulus-secretion coupling of these proteins, protective salivary functions in the oral cavity may be compromised during beta 1-adrenolytic treatment.

Adult↗

PubMed1993

Mutational effects on the cooperativity of Ca2+ binding in calmodulin.

The importance of the aspartate ligand in the +Y Ca2+ coordinating position of two EF-hands of calmodulin has been investigated. Synthetic calmodulin genes were used to produce engineered proteins with the wild-type bovine sequence as well as with aspartate 58 in Ca(2+)-binding site II and/or aspartate 95 in site III changed to asparagine. The macroscopic Ca(2+)-binding constants of the intact calmodulins and of tryptic fragments comprising the N- and C-terminal domains were determined from titrations with Ca2+ in the presence of 5,5'-Br2BAPTA. Substitution of aspartate by asparagine in Ca(2+)-binding site II led to a slight increase in the total free energy change on Ca2+ binding, and the cooperativity of Ca2+ binding to the N-terminal sites was substantially increased. The change from aspartate to asparagine in site III decreased the Ca2+ affinity and also appeared to decrease the positive cooperativity between the sites in the C-terminal domain. Thus, identical mutations in sites II and III were found to result in opposite effects. The data imply that involvement of liganding side chains in interactions other than direct calcium attraction and calcium coordination is of considerable importance for the Ca(2+)-binding process, particularly for the cooperativity.

Allosteric Regulation↗

PubMed1993

Simple reaction-time responses to mechanical and electrical stimuli in human masseter muscle.

The latencies of the simple reaction-time responses for opening and closing of the jaws in response to taps and pushes on teeth and lip shocks were measured in human adults. The reaction times were scored from both the integrated electromyographic (EMG) signal in individual trials, and from the cumulative sum (CUSUM) of the averaged EMG response to 50 stimuli. The mean reaction times for both closing and opening were about 80-90 ms for taps and electrical lip shocks, and about 140 ms for push, measured from the CUSUMs. The reaction times in individual trials were difficult to measure from the EMG signal because of the unsteady baseline. In contrast, the reaction times measured from the CUSUMs were clearly defined by the point at which the record began to move sharply above or below the baseline. However, because of the delays inherent in the CUSUM procedure, they were systematically longer than the means of the trial-by-trial measurements.

Adult↗

PubMed1993

A study of cholinergic and beta-adrenergic components in the regulation of blood flow in the tooth pulp and gingiva in man.

In 10 subjects, laser Doppler flowmetry was used to study whether cholinergic or beta-adrenergic pathways are involved in the control of tooth pulp blood flow (PBF) in response to isometric hand grip and the cold pressor test. We also examined if differences exist between the regulation of blood flow in the tooth pulp and the nearby gingiva (GBF). Isometric hand grip (35% of maximum force, 2 min) and the subsequent ischaemia (2 min) induced a brief rise in PBF and a more long-lasting rise in GBF. Atropine increased heart rate about by 40% and changed the pulpal response to a fall in flow, without altering gingival flow. Propranolol, causing a 20% reduction in heart rate, had no effect on either flow during the actual test, but induced a rise in GBF after the ischaemic period. The cold pressor test (2 min at 0.5 degrees C) resulted in a reduction in PBF and GBF, unaffected by the blocking drugs. With atropine, however, PBF increased immediately after this test. The relative changes in arterial pressure and heart rate were unaffected by the drugs. Our study has demonstrated the existence of cholinergic nervous vasodilation in vessels serving the tooth pulp. Non-adrenergic non-cholinergic mechanisms probably contribute to the evoked rise in GBF during exercise. Beta-adrenoceptors are involved in the control of GBF immediately after isometric exercise. While the two tests under control conditions evoked mostly parallel changes in PBF and GBF, the use of blocking agents showed that blood flow is controlled by different mechanisms in the two adjacent vascular beds.

Adrenergic Fibers↗

PubMed1993

Mechanoreceptors around the tooth evoke inhibitory and excitatory reflexes in the human masseter muscle.

1. The reflex responses evoked in the human masseter muscle by controlled mechanical stimulation of an incisor tooth were examined electromyographically. The stimuli were (slow) pushes and (brisk) taps of about 0.5-3 N peak force, applied orthogonally to the labial surface. 2. The brisk taps elicited a short-latency inhibitory reflex that was often followed by an excitatory peak, as has been described earlier. The inhibition increased as the taps became stronger. 3. Slow pushes evoked a long-latency, primarily excitatory response. The excitation increased with stronger, faster rise-time pushes; however, with the stronger stimuli, the short-latency inhibitory response often became evident before the onset of the excitation. 4. The reflex responses to 3 N pushes and 2 N taps were abolished when the receptors around the tooth were blocked with local anaesthetic, indicating that the response was elicited from receptors located within the periodontal area. 5. Prolonged, rapid-onset displacements evoked a complex reflex response that combined the characteristics of the taps and the pushes. 6. The most likely explanation for the different responses evoked by the pushes and taps is that the patterns of afferent activity elicited by the slow and fast tooth displacements activated different interneuronal pathways to motoneurones. 7. The inhibitory response to taps is essentially a protective reflex which probably serves to reduce the activity of the jaw-closing muscles when one bites unexpectedly on hard objects. It is suggested that the excitatory response may contribute to the muscle activity required to hold food between the teeth during chewing, or may act as a load compensation reflex to control chewing force.

Adolescent↗

PubMed1993

Masseter muscle reflex evoked by tapping on osseointegrated Frialit implants.

Comparison was made of the masseter muscle reflexes evoked by tapping on osseointegrated single-tooth aluminum oxide implants, and on natural teeth in nine patients. Tapping on eight out of nine patients evoked an inhibitory masseter muscle reflex, whereas tapping on all of the natural teeth evoked an inhibitory reflex. The threshold for this reflex was clearly elevated in implants compared to natural teeth. The pathway for the impulses responsible for this reflex and the clinical implications of the elevated threshold are discussed.

Adolescent↗

PubMed1992

Identification of two domains which mediate the binding of activating phospholipids to the plasma-membrane Ca2+ pump.

The stimulation of the purified human erythrocyte calcium pump by acidic phospholipids was investigated using synthetic peptides corresponding to a putative phospholipid-responsive domain [Zvaritch, E., James, P., Vorherr, T., Falchetto, R., Modyanov, N. & Carafoli, E. (1990) Biochemistry 29, 8070-8076] and to the calmodulin-binding domain of the pump. The peptides interfered with the activation of the enzyme by phosphatidylserine and phosphatidic acid in competition assays. The peptide corresponding to the calmodulin-binding domain was found to be the most efficient antagonist. Direct binding measurements using fluorescent derivatives of the peptides confirmed the interaction between the acidic phospholipids and the peptides, and fluorescence titrations of dansylated calmodulin with the purified ATPase showed a direct effect of acidic phospholipids on calmodulin binding. A proteolyzed preparation of the Ca(2+)-ATPase lacking the calmodulin-binding domain confirmed that the phospholipid-induced stimulation is mediated by two sites, one located in the C-terminal portion of the previously identified 44-amino-acid phospholipid-responsive domain, the other in the calmodulin-binding domain.

Amino Acid Sequence↗

PubMed1991

Mutation of the pseudo-EF-hand of calbindin D9k into a normal EF-hand. Biophysical studies.

The two Ca(2+)-binding sites in calbindin D9k, a protein belonging to the calmodulin superfamily of intracellular proteins, have slightly different structure. The C-terminal site (amino acids 54-65) is a normal EF-hand as in the other proteins of the calmodulin superfamily, while the N-terminal site (amino acids 14-27) contains two additional amino acids, one of which is a proline. We have constructed and studied five mutants of calbindin D9k modified in the N-terminal site. In normal EF-hand structures the first amino acid to coordinate calcium is invariantly an Asp. For this reason Ala15, is exchanged by an Asp in all mutants and the mutants also contain various other changes in this site. The mutants have been characterized by 43Ca, 113Cd and 1H NMR and by the determination of the calcium binding constants using absorption chelators. In two of the mutants (one where Ala14 is deleted, Ala15 is replaced by Asp and Pro20 is replaced by Gly, the other where, in addition, Asn21 is deleted), we find that the structure has changed considerably compared to the wild-type calbindin. The NMR results indicate that the calcium coordination has changed to mainly side-chain carboxyls, from being octahedrally coordinated by mainly back-bone carbonyls, and/or that the coordination number has decreased. The N-terminal site has thus been turned into a normal EF-hand, in which the calcium ion is coordinated by side-chain carboxyls. Furthermore, the calcium binding constants of these two mutant proteins are almost as high as in the wild-type calbindin D9k. That is, the extensive alterations in the N-terminal site have not disrupted the calcium binding ability of the proteins.

Amino Acid Sequence↗

PubMed1991

Electrostatic contributions to the binding of Ca2+ in calbindin D9k.

A set of accurate experimental data is provided for Ca2+ ion binding to calbindin D9k, a protein in the calmodulin superfamily of intracellular regulatory proteins. The study comprises both the role of protein surface charges and the effects of added electrolyte. The two macroscopic Ca2(+)-binding constants K1 and K2 are determined for the wild-type and eight mutant calbindins in 0, 0.05, 0.10, and 0.15 M KCl from titrations in the presence of Quin 2 or 5,5'-Br2BAPTA. The mutations involve replacement of surface carboxylates (of Glu17, Asp19, Glu26, and Glu60) with the corresponding amides. It is found that K1K2 may decrease by a factor of up to 2.5 x 10(5) (triple mutant in 0.15 M KCl as compared to the wild-type protein in 0 M KCl). Ca2(+)-binding constants of the individual Ca2+ sites (microscopic binding constants) have also been determined. The positive cooperativity of Ca2+ binding, previously observed at low salt concentration [Linse et al. (1987) Biochemistry 26, 6723-6735], is also present at physiological ionic strength and amounts to 5 kJ.mol-1 at 0.15 M KCl. The electrolyte concentration and some of the mutations are found to affect the cooperativity. 39K NMR studies show that K+ binds weakly to calbindin. Two-dimensional 1H NMR studies show, however, that potassium binding does not change the protein conformation, and the large effect of KCl on the Ca2+ affinity is thus of unspecific nature. Two-dimensional 1H NMR has also been used to assess the structural consequences of the mutations through assignments of the backbone NH and C alpha H resonances of six mutants.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1991

Ca2+ binding in proteins of the calmodulin superfamily: cooperativity, electrostatic contributions and molecular mechanisms.

In a large number of intracellular regulatory proteins of the calmodulin superfamily a pair of closely interacting helix-loop-helix Ca2+ binding sites ('EF hands') constitute the functional unit--an arrangement that enables cooperative binding. We have recently made detailed experimental studies of the binding of Ca2+ ions to calmodulin, its tryptic fragments TR1C and TR2C (which each constitute a globular domain of a pair of EF hands) and calbindin D9k. Macroscopic Ca2+ binding constants have been obtained over a range of ionic strengths (0 to 0.15 M KCl). For calmodulin the measurements indicate that the two separate globular domains TR1C and TR2C retain the Ca2+ binding properties they have in the intact molecule, with positive cooperativity within each domain. The absolute value of the free energy of interaction between the two sites in each domain, a measure of the cooperativity, increases with ionic strength and is greater than or equal to 10 kJ mol-1 at 0.15 M KCl. Two-dimensional 1H NMR studies show that the addition of KCl does not alter the conformation of the protein. In the case of calbindin D9k several categories of mutants have been studied. One group encompasses the effect of protein surface charges 5 to 15 A from the Ca2+ binding sites. Two-dimensional 1H NMR shows that neither the addition of KCl, nor mutations that neutralize the surface charges, change the protein conformation. Although the global structure of calbindin D9k is largely unchanged upon binding of calcium, the structure with only one cation bound is more similar to the (Ca2+)2 form. Interestingly, the dynamical properties of the Ca(2+)-free and the (Ca2+)2-forms of calbindin differ greatly. For example, the rate of NH/ND exchange of the Ca(2+)-free form is on average 200 times faster than that of the (Ca2+)2-form. The results obtained so far point to a non-negligible entropic contribution to the observed cooperativity of Ca2+ binding.

Amides↗

PubMed1991

High-yield purification of HIV-1 proteinase expressed by a synthetic gene in Escherichia coli.

A rapid and simple purification procedure for human immunodeficiency virus type 1 (HIV-1) proteinase from a synthetic gene expressed in Escherichia coli has been developed. The synthetic gene was constructed from oligonucleotides containing several restriction enzyme sites in order to allow simple construction of homologous genes. The protein was translated as a precursor which was autocatalytically processed into the mature protein as shown by N-terminal sequence analysis of the purified protein. Immunoblot analysis was used to verify the nature of the expression product and it was found that 2 of 10 anti-peptide antibodies, covering the whole proteinase sequence, were able to react with the enzyme in crude bacterial lysates. These two anti-peptide antibodies represent a continuous sequence partially overlapping the active site. The purification involves two initial precipitation steps followed by cation-exchange and size-exclusion chromatography. A high yield and a high specific activity were achieved.

Amino Acid Sequence↗

PubMed1991

Force-related changes in the masseter muscle reflex response to tooth-taps in man.

In 14 subjects with no disorder of the masticatory apparatus, excitatory and inhibitory reflex responses in the masseter muscles were derived by standardized mechanical stimulation of the upper central incisor. A series of eight taps was delivered during isometric contraction at 40% of maximum EMG activity, with tapping forces ranging from 0.25-9 N. The mean post-stimulus masseteric EMG complex (PSEC) consisted of up to three inhibitory (I-1, I-2 and I-3) and three excitatory phases. The first inhibitory wave increased in duration with increasing tapping force, and the second and third inhibitory waves occurred at higher thresholds than the first inhibitory wave.

Adolescent↗