Detection and verification of quantitative trait loci for resistance to Dothistroma needle blight in Pinus radiata.

Six related radiata pine ( Pinus radiata) full-sib families were used to detect and independently verify quantitative trait loci (QTLs) for resistance to Dothistroma needle blight, caused by Dothistroma septospora. The detection families had from 26 to 30 individuals each, and had either a common maternal (31053) or paternal (31032) parent; one family (cross 4) consisted of progeny from both parents, 31053 x 31032. Approximately 200 additional progeny from cross 4 were clonally replicated and planted at two sites, with at least five to seven ramets of each individual per site. Marker segregation data were collected from a total of 250 RFLP and microsatellite markers, and single factor ANOVAs were conducted separately for each family and marker. A number of putative associations were observed, some across more than one family. Permutation tests were used to confirm expected probabilities of multiple associations based on chance alone. Seven markers representing at least four QTLs for resistance to Dothistroma were identified as being significant in more than one family; one of these was significant at P<0.05 in three families and highly significant at P<0.01 in a fourth. Further confirmation was obtained by testing those markers that were significant in more than one of the detection families (or highly significant in cross 4) in the clonally replicated progeny from cross 4. Four QTL positions were verified in the clonal populations, with a total percent variation accounted for of 12.5.

QTL associations for density and diameter in Pinus radiata and the potential for marker-aided selection.

A large full-sib family of radiata pine ( Pinus radiata Donn. ex D. Don) was used for quantitative trait locus (QTL) detection and independent verification. QTL detection experiments were carried out for juvenile wood density (JWD) and stem diameter at breast height (DBH) using selective genotyping. Evenly spaced RFLP and microsatellite markers were selected from an existing linkage map. QTLs were verified in an independent set of progeny from the same family. Based on map location, at least eight QTL positions for JWD and two for DBH were detected and verified. The percent variance accounted for by the markers ranged from 0.78% to 3.58%, suggesting a genomic architecture of many genes with small effect. Two unrelated "bridging" families were chosen as candidates for marker-aided selection (MAS), and six microsatellite markers showing an association with JWD or DBH were tested in these families. Of these, four markers showed a consistent association with JWD in one or both of the bridging families. Results from this study provide a basis for MAS in P. radiata.

Electrophysiological properties of electrical synapses between rat sympathetic preganglionic neurones in vitro.

1. The electrophysiological properties of electrical synaptic transmission between sympathetic preganglionic neurones (SPNs) in slices of rat spinal cord were investigated using simultaneous dual-electrode patch-clamp recordings. Electrotonic coupling was directly demonstrated between 21 pairs of SPNs. 2. Coupling coefficients determined from the steady-state response of both neurones to current steps injected into either neurone ranged from 0. 02 to 0.48 (0.18 +/- 0.02, mean +/- s.e.m.). Synapses were bidirectional and symmetrical for the majority of connections with coupling coefficients similar in either direction. Asymmetrical coupling between a minority of cell pairs was due to differences in passive neuronal properties rather than rectification of the synaptic conductances. 3. Action potentials were manifest in adjoining cells as biphasic electrical postsynaptic potentials (ePSPs), composed of a rapid depolarising component followed by a more prolonged hyperpolarisation with amplitudes of 1.2 +/- 0.2 and 2.1 +/- 0.6 mV, respectively. 4. Postsynaptic potentials resembled low-pass filtered presynaptic spikes with frequency dependence determined by the junctional conductance and postsynaptic membrane properties. Increases in presynaptic action potential frequency caused attenuation of the hyperpolarising component of the ePSP that was attributed to shorter duration presynaptic spikes being more markedly filtered. 5. Synchronisation of spontaneous action potentials between electrotonically coupled neurones was driven by subthreshold membrane potential activity resembling repetitive ePSPs. Synchronous spike firing in previously silent neurones could be driven by suprathreshold ePSPs induced by suprathreshold depolarisation of a single adjoining neurone. 6. These data characterise reliable communication of sub- and suprathreshold activity by electrical synapses enabling synchronised SPN firing which may contribute to generation of coherent sympathetic rhythms and promote summation of inputs to postganglionic neurones.

Metabotropic glutamate receptor-mediated excitation and inhibition of sympathetic preganglionic neurones.

The effects of metabotropic glutamate receptor (mGluR) subtype selective compounds on the excitability of sympathetic preganglionic neurones (SPNs) were investigated. Non-selective mGluR agonists (1S,3R)-aminocyclopentane-1,3-dicarboxylic acid and (2S,1'S,2'S)-2-(carboxycyclopropyl)glycine, induced dose-dependent depolarisations in 96 and 75% of SPNs, respectively and hyperpolarisations in 2 and 21% of SPNs. Both agonists could induce subthreshold membrane potential oscillations in previously non-oscillating SPNs and either increased or reduced the frequency of spontaneously occurring oscillations. A selective group I mGluR agonist, 3,5-dihydroxyphenylglycine, depolarised all SPNs tested, induced oscillations in membrane potential of otherwise non-oscillating SPNs and increased the frequency of spontaneous oscillations. Agonists with selectivity for group II mGluRs (1S,3S)-aminocyclopentane-1,3-dicarboxylic acid and (S)-4-carboxy-3-hydroxy-phenylglycine ((S)-4C3HPG) did not induce depolarising responses. However (S)-4C3HPG induced hyperpolarising responses associated with a reduction in the frequency of spontaneous oscillations in two of six SPNs tested. Depolarising and hyperpolarising responses were maintained in the presence of tetrodotoxin indicating a direct action of the agonists upon SPNs. In individual SPNs responses of opposite polarity could be induced from the same initial membrane potential using different agonists, indicating that the opposing responses involved different ionic mechanisms. The broad spectrum mGluR antagonist (S)-alpha-methyl-4-carboxyphenylglycine and the selective group I mGluR antagonist (S)-4-carboxyphenylglycine reversibly depressed mGluR agonist induced depolarisations. These results indicate that SPNs express two mGluR populations with opposing actions on neuronal excitability: group I mGluRs depolarise SPNs and can drive oscillatory membrane potential activity; a minority of SPNs express group II mGluRs which mediate membrane hyperpolarisations and reduce the frequency of membrane potential oscillations.

Actions of the anaesthetic Saffan on rat sympathetic preganglionic neurones in vitro.

1. Whole-cell patch-clamp recordings were used to investigate the effects of the anaesthetic Saffan on the electrophysiological properties of sympathetic preganglionic neurones (SPNs) in rat spinal cord slices. 2. Saffan (1-54 microM) abolished or reduced the frequency of spontaneous action potential firing and abolished spontaneous, sub-threshold membrane potential oscillations. Saffan caused dose-dependent decreases in input resistance and depending upon the initial resting membrane potential, either a depolarization, a hyperpolarization or no change in membrane potential. 3. Responses to Saffan were blocked by the GABAA receptor antagonists bicuculline (5-20 microM) and picrotoxin (20 microM), but not by the glycine receptor antagonist strychnine (20 microM) indicating that they were mediated by GABAA receptors. 4. Changes in the properties of SPN action potentials were also observed. In the presence of Saffan the amplitude and duration of the action potential after-hyperpolarization were reduced and larger depolarizations were required in order to evoke trains of action potentials. 5. To examine the effects of Saffan on electrotonic coupling between SPNs, experiments were performed with the Na+ channel blocker QX-314 in the intracellular solution and antidromic oscillations were evoked by ventral root stimulation. Saffan failed to abolish antidromic oscillations, but reduced their amplitude and duration. This indicates that the abolition of spontaneous membrane potential oscillations was not a direct effect on the coupling between SPNs, but was a result of the abolition of spontaneous activity by Saffan. 6. The responses to Saffan occurred within the plasma concentration range of Saffan during anaesthesia, suggesting that the electrophysiological properties of SPNs may be altered during anaesthesia with Saffan. This would be expected to lead to changes in sympathetic tone and in the integration of sympathetic output.

Electrotonic coupling between rat sympathetic preganglionic neurones in vitro.

1. Using the whole-cell recording technique in rat spinal cord slices we have shown that 26% of sympathetic preganglionic neurones (SPNs) show spontaneous membrane potential oscillations. These oscillations consist of trains of biphasic waves, which we have termed spikelets because of their similarity to truncated action potentials. 2. The spikelets were inhibited by TTX and anaesthetics such as alpha-chloralose but not by the intracellular application of lidocaine N-ethyl bromide (QX-314). 3. By stimulating the ventral roots we have demonstrated the presence of short-latency depolarizations (SLDs) in oscillating neurones. These SLDs have a similar waveform to the spontaneous spikelets, and also show the ability to override the frequency of occurrence of the spontaneous spikelets. These observations suggest that the spikelets result from electrotonic coupling between the oscillating SPNs. 4. SLDs were also observed in a population of non-oscillating, electrotonically coupled, quiescent SPNs. It was possible to induce oscillations in these neurones by the injection of depolarizing current (in the presence of QX-314), suggesting that these neurones are also gap-junction coupled. 5. Simultaneous whole-cell recordings were obtained from twenty-three pairs of SPNs. Two pairs displayed both spontaneous, synchronized oscillations and action potentials. Electrotonic coupling was confirmed by the detection of membrane polarization in both neurones in response to current injected into one neurone. In a further two pairs of quiescent SPNs, injection of depolarizing current pulses into one neurone induced action potential discharge in that neurone and a depolarization and oscillations in the other neurone. 6. The ability of groups of electrotonically coupled SPNs to generate spontaneous discharges within the spinal cord provides a novel mechanism for the integration and synchronization of information within the sympathetic nervous system.

Patient-controlled analgesia: a method for the controlled self-administration of opioid pain medications.

The purposes of this article are to introduce the reader to patient-controlled analgesia (PCA) and to summarize its use in several selected pain-related conditions. patient-controlled analgesia is a relatively new technique for managing pain in which patients are able to self-administer small doses of opioid analgesic medications when needed. The authors briefly review some of the problems associated with current and previous opioid delivery strategies and highlight the advantages of PCA over these other methods. They then discuss the components of the PCA system and briefly describe how the system is operated and controlled. In this discussion, the authors indicate an appropriate therapeutic goal and suggest knowledge requirements for the effective use of PCA. The authors close with a brief summary of several reports describing the use of PCA in the management of postoperative pain, cancer pain, and pain associated with labor and delivery. Indications and contraindications for use in these conditions are presented. Because physical therapists often play a major role in pain management, it is important for them to be well informed with regard to recent developments in this rapidly developing area of clinical practice.

Failure of transcutaneous electrical nerve stimulation in the conventional and burst modes to alter digital skin temperature.

The purpose of this study was to determine if transcutaneous electrical nerve stimulation (TENS), when applied using stimulation settings that might be used in clinical practice, produces alterations in skin temperature. Stimulation in the conventional and burst modes was applied to the skin of the forearm overlying the median nerve in a healthy human subject. Skin temperature was measured at the index finger, little finger, and on the volar surface of the forearm in the stimulated limb. TENS applied for 20 minutes in the conventional mode at an intensity sufficient to produce a perceptible though not uncomfortable sensation and a slight muscle contraction of the forearm musculature failed to alter skin temperature at any of the three measurement sites. Stimulation using the same pulse characteristics delivered in the burst mode failed to alter skin temperature of the fingers but produced a significant increase in skin temperature at the forearm. The increase in skin temperature at the forearm persisted for four minutes following the cessation of stimulation after which skin temperature returned to prestimulation levels. We conclude that TENS, as applied in this study, does not influence skin temperature in the peripheral distribution of the nerve stimulated, and hypothesize that the rise in skin temperature at the forearm was a result of increased blood flow in the forearm muscles produced by the stimulation-induced contraction of those muscles.

Pain measurement. An overview of two commonly used methods.

Two pain-measurement tools in particular have gained widespread acceptance by clinicians who work in the area of pain management: the visual analogue scale and the McGill Pain Questionnaire. The purpose of this paper is to describe briefly both of these methods and to consider some of their strengths, weaknesses, and applications.

Conductive differences in electrodes used with transcutaneous electrical nerve stimulation devices.

The purpose of this study was to document conductive differences among commercially available electrodes used with transcutaneous electrical nerve stimulation (TENS) devices. Impedance within a model system involving a human subject was calculated from oscilloscopic tracings of the pulse waveform for each of 25 different electrode types. Impedance values ranged from 1,000 to 7,800 omega Possible reasons for these differences are discussed. The observation that electrodes vary in their impedance and can thereby affect the stimulus applied to the skin raises the question of whether electrode choice might affect the clinical effectiveness of TENS. Attention is drawn to the skin electrodes as a variable that may affect the results of clinical and basic studies involving TENS.

Selected problems in the use of transcutaneous electrical nerve stimulation for pain control--an appraisal with proposed solutions. A special communication.

The purposes of this communication are 1) to call attention to several problematic issues related to the use of transcutaneous electrical nerve stimulation for pain control and 2) to consider strategies for correcting existing problems and averting additional problems in the future. Specific problems within two general areas of difficulty are identified, and proposed solutions to these problems are presented. It is hoped that discussion of these issues will encourage comment and debate, bring about further objective and critical evaluation of the effectiveness and value of TENS for pain control, and lead to a clearer understanding of the responsibilities of manufacturers and users of this therapeutic modality.

Transcutaneous electrical nerve stimulation for pain control after spinal fusion with Harrington rods. A clinical report.

During the past two years, we have been using transcutaneous electrical nerve stimulation (TENS) for pain control after spinal fusion with Harrington rods. This report describes our postoperative pain management program and our evaluation of its effectiveness. We examined postoperative pain medication usage and found that patients using TENS received fewer doses of several commonly prescribed pain drugs than did patients not using TENS devices. Difficulties we encountered in evaluating our program are described along with recommendations for other researchers interested in assessing TENS in the management of postoperative pain.

Quantitative measure of cutaneous sensation. Two-point discrimination values for the face and trunk.

This study was performed to assess two-point discrimination ability for skin areas of the face and trunk. Using a compass-type instrument, I determined two-point discrimination values for three areas on the face and eight, nonoverlapping regions of the neck and trunk in a sample of 43 healthy young adult men and women. Mean values for the face ranged from 14.9 mm over the eyebrow to 10.4 mm along the lateral aspect of the mandible. Values for the neck and trunk ranged from 35.2 mm for skin of the lateral neck to 55.4 mm for the region immediately lateral to the C7 spinous process. I also found interindividual variation in two-point discrimination ability for a given skin area. Except for skin overlying the body of the mandible, where values for women were lower than those measured in men, no significant differences in discrimination ability were found between men and women. Although assessment of two-point discrimination is useful in the clinical evaluation of certain types of patients, the existence of intraindividual and interindividual differences suggests that therapists must interpret the results of these tests with caution.

Primary afferent projections to the sacral secondary visceral gray: a quantitative electron microscopic examination in the cat.

Afferent input to the sacral secondary visceral gray matter was studied by quantitatively assessing synaptic degeneration that follows ipsilateral sacral dorsal rhizotomy. Five days after the lesions, slightly more than 25% of the synaptic contacts showed degenerative changes. The majority of the reactive synapses contained clear spherical vesicles (S type). Boutons containing dense core vesicles (GS type) also showed degenerative changes, and together with S boutons constituted more than 96% of the reactive terminals. Degenerating S type synapses were found only on dendrites whereas GS boutons were observed on both cell bodies and dendrites. Further examination revealed that although there were more S than GS synapses in the nucleus, the percentage of GS terminals affected was greater than that for S type boutons. These results demonstrate a direct primary afferent input to the sacral secondary visceral gray matter utilizing chiefly two different synaptic types.

Limits of two-point discrimination ability in the lower limb in young adult men and women.

The purpose of this study was to determine the limits of two-point discrimination ability in the lower limb in a sample of healthy, young adult men and women. Eleven different areas of skin were tested with a commercially available compass-type instrument to determine the shortest distance at which two simultaneously applied, nonpainful, light-touch stimuli could be perceived. Mean two-point discrimination values in the lower limb ranged from 43.6 mm for skin overlying the medial surface of the leg to 6.6 mm on the plantar surface of the tip of the great toe. The results also demonstrated interindividual variation in two-point discrimination for a given area of skin. In the majority of skin areas tested, mean values obtained from men and women were essentially the same. In several of the test areas, however, women were significantly better than men in their two-point discrimination ability. The observed intraindividual and interindividual variation suggests that although two-point testing is a useful clinical tool, caution is necessary when interpreting the results from patients with alterations in cutaneous sensibility.