A novel size-selective airborne particle size fractionating instrument for health risk evaluation.

Health risks associated with the inhalation of airborne particles are known to be influenced by particle size. Studies have shown that certain nanoparticles, with diameters <100 nm, have increased toxicity relative to larger particles of the same substance. A reliable, size-resolving sampler able to collect a wide range of particle sizes, including particles with sizes in the nanometre range, would be beneficial in investigating health risks associated with the inhalation of airborne particles. A review of current aerosol samplers used for size-resolved collection of airborne particles highlighted a number of limitations. These could be overcome by combining an inertial deposition impactor with a diffusion collector in a single device. Verified theories of diffusion and inertial deposition suggested an optimal design and operational regime. The instrument was designed for analysing mass distribution functions. Calibration was carried out using a number of recognized techniques. The sampler was tested in the field by collecting size-resolved samples of lead containing aerosols present at workplaces in factories producing crystal glass. The mass deposited on each screen proved sufficient to be detected and measured by an appropriate analytical technique. Mass concentration distribution functions of lead were produced. The nanofraction of lead in air varied from 10 to 70% by weight of total lead.

The effects upon the activity of hand and forearm muscles of intracortical stimulation in the vicinity of corticomotor neurones in the conscious monkey.

Corticomotor (CM) neurones were identified in three conscious macaque monkeys by the presence of post-spike facilitation (PSF) in spike-triggered averages of e.m.g. recorded from intrinsic hand and forearm muscles during performance of a precision grip task. Post-spike effects were compared with those produced by single-pulse intracortical microstimulation (ICMS), with strengths of 4-20 microA, delivered at the site of 47 CM cells. Most muscles facilitated by a CM cell were also facilitated by ICMS at the site of the cell. ICMS effects were stronger: at 10 microA, the amplitude of ICMS-evoked facilitation was on average 2.8 times greater than PSF, and 6.9 times greater at 20 microA. Onset latency of ICMS-evoked facilitation was consistently longer (by 1.7 and 1.3 ms at 10 and 20 microA respectively) than PSF, and it is suggested that this results from the indirect, trans-synaptic excitation of CM cells by ICMS. Post-spike suppression was rarely seen (7/421 compared to 105/421 cases of PSF). In contrast, suppression and facilitation were equally common in response to ICMS. The synaptic mechanisms underlying these effects were explored in 5 anaesthetised macaque monkeys. ICMS facilitated a greater proportion of the tested muscles than did the CM cell recorded at the stimulus site. The results suggest the juxtaposition in the motor cortex of CM neurones with different 'muscle fields'. The merits of STA and ICMS for exploring cortical organisation are discussed.

Selective facilitation of different hand muscles by single corticospinal neurones in the conscious monkey.

1. Post-spike facilitation of e.m.g. activity by monkey motor cortex neurones has been investigated in different hand and forearm muscles. 2. Seventy-eight neurones were recorded concurrently with between five and ten different muscles. Forty-seven neurones were identified as cortico-motor by the presence of post-spike facilitation in the spike-triggered average of at least one of the tested muscles. 3. All forty-seven cortico-motor neurones showed clear increases in activity during performance of a precision grip task by the monkey, and all of them were co-activated with the sampled muscles. 4. To assess the divergence of facilitation from a single cortico-motor neurone to different muscles, spike-triggered averages were constructed with all of the concurrently recorded muscles. The number of muscles in the sample, and the number of muscles showing post-spike facilitation, were corrected by excluding any post-spike facilitation which could have arisen by cross-talk between the different pairs of e.m.g. electrodes. 5. Most cortico-motor neurones produced post-spike facilitation in a restricted number of tested muscles. The mean number of post-spike facilitation-bearing muscles per cortico-motor cell rose from 1.4 +/- 0.5 (S.D.) when five muscles were sampled to 2.0 +/- 1.5 when ten were sampled. On average, each cortico-motor neurone produced post-spike facilitation in 27% of the tested muscles. Only three of forty-seven cortico-motor neurones gave post-spike facilitation in half or more of the tested muscles. 6. The distribution pattern of post-spike facilitation among the muscles sampled with a given cortico-motor neurone was not altered when the spike-triggered averages were constructed from cortico-motor cell and e.m.g. activity recorded during two different phases of the precision grip task, or during performance of a quite different, power grip, task. 7. Cortico-motor cells which produced post-spike facilitation in two or more different muscles often did so in muscles with synergistic functions. 8. It is suggested that cortico-motor neurones may contribute to relatively independent finger movements by virtue of their selective facilitation of hand muscles leading to a fractionated pattern of muscle activity.

Corticospinal facilitation of hand muscles during voluntary movement in the conscious monkey.

1. The method of spike-triggered averaging has been used to detect a direct influence of pyramidal tract neurones on the activity of hand and forearm muscles in conscious monkeys trained to perform repetitive movements of the hand and fingers. Gross electromyograms (e.m.g.s) from individual muscles were rectified and synchronously averaged with respect to the discharge of single, antidromically identified pyramidal tract cells in the 'hand' area of the pre-central gyrus. 2. The presence in an average of a post-spike facilitation which could be revealed reproducibly from successive epochs of recording and was clearly larger than the biggest fluctuations seen in pseudo-randomly triggered averages of the same e.m.g. data, was taken to indicate a direct cortico-motoneuronal excitatory influence. 3. 55% of cortical neurones analysed showed post-spike facilitation in one or more recorded muscle and 7% showed post-spike suppression. In terms of the total number of muscle-neurone combinations analysed, the proportions showing post-spike effects were 18 and 1% respectively. These figures have been influenced by the pre-selection of neurones for analysis according to restrictive criteria. The neurones selected (a) were recorded at cortical loci where weak intracortical microstimulation could evoke finger movements, (b) could be activated antidromically at short latency by medullary pyramidal tract stimulation, (c) showed natural discharge activity which was clearly modulated in relation to voluntary finger movements, and (d) were located in the anterior bank of the central sulcus. The results provide some evidence to vindicate these criteria. 4. The strongest post-spike facilitation observed had a peak which was 42% higher than the average pre-spike level of e.m.g. activity, but most were within the range 5-20%. Facilitation peaks below about 3% could not have been resolved from the 'noise' in the averages. The mean latency from cell discharge in the cortex to the start of the post-spike facilitation was 11.2 ms (range 7.4-17.2) for intrinsic hand muscles and 9.8 ms (range 4.1-15.0) for forearm muscles. These latencies were compared with the latencies of responses to intracortical microstimulation and to stimulation of the medullary pyramidal tract. 5. Evidence was obtained suggesting that the latency for cortico-motoneuronal activation of an individual motor unit was commonly subject to considerable variability and that different motor units of a muscle could be facilitated by the one cortical neurone at different latencies. These factors are thought to contribute to an elongation of the time course of post-spike facilitation.(ABSTRACT TRUNCATED AT 400 WORDS)

Antidromic excitation of motoneurons by intramuscular electrical stimulation.

The terminal branches of motoneurons can be excited by electrical current pulses delivered via needles inserted intramuscularly. Observations are reported which indicate that tungsten semi-microelectrodes suitably positioned in a muscle belly can excite antidromically a large proportion of the alpha-motoneurons of that muscle, at stimulus strengths weak enough to avoid spread of excitation to motor units of adjacent muscles. Some gamma-motoneurons and afferent axons can also be excited by intramuscular stimulation. This technique of antidromic excitation will find application in electrophysiological studies on motoneurons of small muscles, such as intrinsic hand and foot musculature, where the individual muscle nerves are too fragile to be dissected and mounted on stimulating electrodes in the conventional manner.

Response of soleus Ia afferents to vibration in the presence of the tonic vibration reflex in the decerebrate cat.

1. Micro-electrode recordings were made from single Ia afferents in the intact nerve to the soleus muscle in the decerebrate cat while the muscle was developing a tonic vibration reflex. This was done in order to test how effectively the afferents were excited by the vibration, and to see if any insecurity in driving might be related to tremor.2. When the amplitude of vibration was 50 mum, and the tonic vibration reflex was reasonably well developed (> 1 N of active tension) all but one of forty-four Ia afferents were driven 1:1 by the vibration. Most were still driven by 30 mum vibration. The vibration, consisting of a train of discrete pulses at 150 Hz, was applied longitudinally in combination with a stretch of 1 mm to make the muscle taut.3. If the reflex was poorly developed (active tension < 1 N) the driving was on average less secure. However, fourteen of eighteen afferents then studied were still driven 1:1 by 50 mum vibration. The lower level of excitation by vibration was thought to be due to a deficiency of spontaneous fusimotor activity, because stroking the cat's tail or other similar gentle manipulation led each of the three misbehaving afferents so tested to be driven securely by 50 mum vibration; at the same time the reflex tension increased.4. Additional, indirect evidence favouring widespread security of Ia driving by 50 mum vibration in the presence of the reflex was obtained by modulating the amplitude of the 150 Hz vibration with a 7-10 Hz square wave and detecting any tension fluctuations at that frequency by spectral analysis. Small degrees of modulation (e.g. < 10%) produced little if any effect, although larger depths of modulation had a powerful action.5. When the amplitude of vibration was reduced to permit insecure driving but still to elicit a reflex response, the fluctuations in Ia firing pattern were unlike those previously seen in the de-efferented muscle. Spectral analysis showed that these firing fluctuations bore a general similarity to the tremor in the same preparation, but measurement of coherence demonstrated that the tremor and Ia firing were not well related. This was probably because individual Ia afferents were primarily influenced by local factors, and provides further evidence against the tremor of this preparation being attributable to the action of the stretch reflex.

Motor unit firing and its relation to tremor in the tonic vibration reflex of the decerebrate cat.

1. The discharge of single motor units has been recorded from the soleus muscle of the decerebrate cat during the tonic vibration reflex elicited isometrically, to further understanding of the tremor that is seen in the reflex contraction. The reflex was elicited by pulses of vibration of 50 micrometers amplitude at 150 Hz, and up to four units were studied concurrently. 2. Individual units fired rather regularly and at a low frequency (range 4-14 Hz). The rate of firing of any unit normally fell within the frequency band of the tremor recorded at the same time. On comparing different preparations a higher frequency of tremor was associated with a higher frequency of motor firing. 3. The responses of pairs of motor units recorded concurrently during repeated production of the reflex were compared by cross-correlation analysis; over 1000 spikes from each train were normally used for this. The major of the cross-correlograms were flat with no overt sign of any synchronization between the units other than that due to the vibration. 4. Clear indications of correlated motor unit firing could be produced deliberately by modulating the amplitude of vibration at a frequency comparable to that of the normal tremor and thereby introducing a rhythmic component into the tonic vibration reflex. 5. About 20% of the cross-correlograms obtained during normal tremor showed varying amounts of an irregular 'waviness' suggesting a possible correlation between the times of firing of a pair of units. But such waves never developed steadily throughout the period of analysis, in contrast to the comparable waves produced on modulating the vibration. Similar waves were seen on cross-correlating a motor unit with an electronic oscillator, confirming that their occurrence does not necessarily demonstrate the existence of active neural interactions. 6. It is concluded that there is no strong and widespread neural synchronizing mechanism active during the tonic vibration reflex, although the possibility of some weak neural interactions has not been excluded. The findings favour the idea that the tremor in this preparation is simply the inevitable result of motor units discharging asynchronously, but at closely similar subtetanic frequencies.

Divergent collaterals from deep cerebellar neurons to thalamus and tectum, and to medulla oblongata and spinal cord: retrograde fluorescent and electrophysiological studies.

In cat the existence of collaterals from deep cerebellar neurons, which project to mesencephalon and thalamus has been investigated anatomically by means of the multiple retrograde fluorescent tracer technique as well as electrophysiologically by means of conventional antidromic techniques. Both sets of data indicate that several neurons in the medial nucleus, which project to mesencephalon and thalamus, also distribute collaterals to medulla oblongata and spinal cord. These branching neurons were principally located in the caudal and intermediate portions of the medial nucleus. The electrophysiological data in addition indicate that the branching point of the neurons in the medial nucleus is located relatively close to the cell soma. The anatomical findings show a further group of branching neurons in the lateral nucleus at the border with the interpositus nuclei. The majority of these latter neurons distribute collaterals to medulla oblongata but some distribute collaterals to spinal cord. However, it could not be decided as yet whether the collaterals to the medulla oblongata terminate either in medullary medial reticular formation or in inferior olive or in both.

Comparison of electromyogram spectra with force spectra during human elbow tremor.

1. The tremor that develops when the elbow is flexed against a spring attached at the wrist has been analysed by determining the 'power' spectrum of the demodulated surface e.m.g. recorded from two of the active muscles, biceps and brachioradialis. This was compared with the corresponding force spectrum obtained by analysis of the force developed at the fixed end of the spring (normally one of stiffness 2.8 N/mm); this force is directly proportional to the movement at the wrist. 2. When the subject was maintaining a high target force (100-160 N), with the aid of a visual display, the tremor was large with a large sharply tuned peak in the force spectrum and there was a clear peak at the same frequency in the e.m.g. spectrum. The coherence (gamma) between the force peak and the corresponding e.m.g. peak typically had a value of 0.95 or above, indicating a high degree of correlation. 3. On developing the same target force against a rigid restraint (70 N/mm) the peak in the force spectrum was absent or very much smaller and less sharply tuned. More particularly, the tremor-related peak seen in the e.m.g. spectrum under compliant conditions was no longer present under rigid conditions. 4. At low target forces (20-40 N) with compliant loading there was a small peak in the force spectrum but no peak could be detected in the e.m.g. spectrum. With increasing target force the mechanical tremor increased considerably and a peak progressively emerged from above the background level in the e.m.g. spectrum, accompanied by the development of a corresponding peak in the coherence spectrum. Thus the difference in detectability of peaks in the e.m.g. and force spectra might simply result from differences in the background 'noise' level of the two types of spectra. 5. Changing the spring stiffness in the range 0.7-12.5 N/mm altered the frequency of the mechanically recorded tremor by 1.5-2 Hz and the peak in the e.m.g. spectrum shifted in approximate correspondence. 6. The findings support the view that the tremor seen with compliant loading of the arm is due to the stretch reflex. In addition, this work should help define the conditions under which spectral analysis of the gross e.m.g. is of practical utility.

Enhancement by agonist or antagonist muscle vibration of tremor at the elastically loaded human elbow.

1. Human subjects attempted to maintain a constant force by flexing their elbow against a spring which was attached to a force transducer at one end and the subject's wrist at the other. The tremor at 8-12 Hz which develops in this situation was enhanced in amplitude with negligible change of frequency by applying vibration at 100 Hz to the tendon either of an agonist muscle (biceps brachii) or of the antagonist (triceps brachii). The tremor was assessed by spectral analysis of the force records and measurement of the peaks in the spectra. The compliance of the spring was normally 2.8 N/mm and target forces of 40-120 N were used. 2. The percentage increase in the tremor on applying vibration was relatively independent of target force, although the absolute amounts of tremor increased markedly with increasing target force. The average increase was greater for vibration of triceps than for biceps (70% as opposed to 37%, averaged between subjects and over a range of forces). 3. When the spring was replaced by a rigid connexion there was usually no clear tremor peak either in the presence or absence of vibration. Vibration, however, tended to increase the general noisiness of the force signal. 4. Qualitatively similar effects were seen when the elbow exerted an extending force so that triceps became the agonist and biceps the antagonist. 5. The tremor peak present in the spectrum of the demodulated electromyogram during vigorous tremor increased in size when vibration made the tremor larger. 6. The effect of a rhythmic afferent input was studied by modulating the amplitude of the vibration at 8-9 Hz, to correspond to the tremor frequency, while the subject pulled against a rigid attachment. Both the e.m.g. and the tension spectra contained peaks at the modulation frequency. The raw force records showed that, with reference to the modulation, the effects of biceps and of triceps vibration were approximately 180 degrees out of phase with each other, as would occur if vibration of one were having an excitatory action, and vibration of the other an inhibitory action. Moreover, in each case the effect on force (whether excitatory or inhibitory) lagged about half a cycle on the vibration envelope, as required for such reflexes to help in the generation of tremor. 7. It is suggested that vibration increases the modulation of Ia firing elicited by a given movement tremor and this, by means of the stretch reflex arc, enhanced the tremor. The powerful action of vibration of the antagonist illustrates, it would seem, the functional effectiveness under normal conditions of a spinal inhibitory pathway, most probably the Ia disynaptic route. The findings are also discussed in relation to the increase in stretch reflex gain that occurs in association with increasing strength of voluntary contraction.

Tremor in the tension developed isometrically by soleus during the tonic vibration reflex in the decerebrate cat.

1. Irregularities in the development of tension during the tonic vibration reflex of the soleus muscle of the decerebrate cat have been analysed into their frequency components. The reflex was recorded isometrically and elicited by longitudinal vibration, normally at 150 Hz. The amplitude of vibration was set so as to elicit a maximal reflex response, suggesting 1:1 driving of the majority of the Ia afferents at the frequency of vibration. 2. The resulting power spectrum regularly showed a well marked tremor peak separated by a trough from any slow irregularities. The predominant frequency of this tremor varied from 4 to 11 Hz in different preparations, with a mean of 7.4 Hz; on average, frequencies within 1.7 Hz on either side contained over half the power of the predominant frequency. Altering the frequency of vibration did not alter the distribution of tremor frequencies. 3. The root mean square value of the tension irregularities, over the range 4-14 Hz, varied from 12 to 110 mN in different preparations (median value, 23 mN); this was superimposed on mean active reflex tensions varying from 2 to 10 N. 4. The 'tremor' due to a single motor unit was estimated from spectral analysis of tetanic contractions of the whole muscle and decreased with increasing frequency of activation. Comparison of the single unit values with the tremor seen during vibration in the same preparations showed that equivalent amounts of tremor to the latter could typically have been produced by the continued synchronous contraction of about five 'average' motor units firing at the predominant tremor frequency. 5. When a tonic stretch reflex was present its tremor frequencies did not differ consistently from those of the tonic vibration reflex. On average, the tremor was smaller for the stretch reflex than for the tonic vibration reflex; the difference was usually slight and might have been related to the stretch refex tension being smaller. 6. Evidence was obtained that the tremor was not due to any insecurity of 1:1 driving of the Ia afferents by the vibration. First, the tremor did not increase when the amplitude of vibration was decreased sufficiently to ensure that the degree of 1:1 driving must have been reduced. Secondly, the introduction of a comparable 'artificial tremor' by sinusoidally oscillating the muscle at low frequency did not produce the e.m.g. response that would have been expected if the applied 'tremor' had been modulating the firing of the Ia or any other group of afferents. 7. It is concluded that the observed tremor cannot be attributed to 'oscillation in the stretch reflex arc', though without prejudice to the role of this mechanism under other conditions and especially when the recording is not isometric. However, the genesis of the tremor has not been established and much of it might result simply from the chance synchronization of motor units that are firing below their tetanic fusion frequency.

The effect of a preceding stimulus on temporal facilitation at corticomotoneuronal synapses.

1. Intracellular recordings were made of minimal corticomotoneuronal e.p.s.p.s in lumbar motoneurones of anaesthetized monkeys. For intervals of 2 msec and greater between paired cortical shocks, the average time course of facilitation of the second e.p.s.p. with respect to the first could be fitted closely by a negative exponential with a time constant of 10 msec.2. In the same motoneurones, ;triplets' of corticomotoneuronal e.p.s.p.s were generated by delivering three identical stimuli to the motor cortex. Considering the triplet as a conditioning e.p.s.p. followed by a test pair, the facilitation of the third e.p.s.p. with respect to the second was measured for various combinations of test and conditioning intervals. In each case the amplitude of the third e.p.s.p. was also compared with that of the first (conditioning) e.p.s.p.3. The effect of a brief conditioning interval was to reduce considerably the facilitation of the third e.p.s.p. with respect to the second at all test intervals from 2 to 50 msec. Combinations of brief conditioning intervals (e.g. 2 or 5 msec) and long test intervals (e.g. 20 or 50 msec) caused the third e.p.s.p. to be smaller than the second. As the conditioning interval lengthened, facilitation in the test pair increased towards the unconditioned values at all test intervals.4. Facilitation of the third e.p.s.p. with respect to the first could be described approximately as the linear addition of two facilitation components, one due to the conditioning input and one due to the first stimulus of the test pair. Each component followed the same negative exponential time course as found for an isolated pair of e.p.s.p.s and each of the first two inputs contributed to the facilitation of the third e.p.s.p. as if the other of these two inputs had not occurred.