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A H Pullen

Publications and source records attributed to A H Pullen.

At least 19 recordsLinked to original sources
PubMed1995

Diversity in localisation of nitric oxide synthase antigen and NADPH-diaphorase histochemical staining in sacral somatic motor neurones of the cat.

Nitric oxide synthase (NOS) immunoreactivity occurred in about 60% of ventromedial, ventrolateral, and sphincteric motoneurones in cat sacral spinal cord. Proportions of sacral motoneurones histochemically stained for NADPH-diaphorase, were similar, and equivalent in size to those immunoreactive for NOS, suggesting co-localisation of diaphorase and NOS. Double staining techniques revealed that NOS co-localised with NADPH-diaphorase in approximately 60% of sacral motoneurones. Some remaining motoneurones exhibited neither NOS nor NADPH-diaphorase, while others exhibited only NOS or NADPH-diaphorase, indicating phenotypic differences among sacral motoneurones.

Animals↗
PubMed1995

Phosphorylated neurofilament antigen redistribution in intercostal nerve subsequent to retrograde axonal transport of diphtheria toxin.

A novel enzyme-linked immunosorbent assay technique using specific monoclonal antibodies has been used to examine the proximal-distal distribution of phosphorylated neurofilament proteins (pNF) in normal feline intercostal nerve, and to compare it with that following retrograde axonal transport of the ADP-ribosylating protein diphtheria toxin (DTX) to thoracic motoneurones. The molecular target of DTX is elongation factor 2 which resides solely in the cell body. Normal intercostal nerves exhibited significantly higher amounts of the 200-kDa pNF-H, 160-kDA pNF-M, and 68-kDA pNF-L in proximal nerve than in the distal nerve. The overall content of all three triplet pNF proteins decreased 3 days after injection of DTX, but the normal proximal-distal gradient was retained. By 8 days post DTX injection, the proximal-distal gradient had reversed, with proximal nerve starved of pNF-H and pNF-M and distal nerve showing abnormally high pNF-L content. Correlative immunocytochemistry of spinal cords from normal animals verified that pNF-H and pNF-M are confined to efferent axons in the spinal grey matter, and that motoneurones are only reactive for pNF-L. At 8 days following toxin treatment, motoneurones in the ipsilateral ventral horn were strongly immunoreactive for all pNF. Contralateral motoneurones were non-reactive. Onset of abnormal perikaryal pNF immunoreactivity at 3 days precedes onset of ultrastructural cytopathology. Together these results indicate an early deficit in transference to the axon of NF proteins synthesised prior to full toxicity, probably because of a toxin-induced failure in regulation of phosphorylation-dependent NF assembly and turnover immediately prior to entry into the proximal axon. Results are discussed in relation to diphtheritic motoneuronopathy.

Animals↗
PubMed1995

Ultrastructural abnormalities with inclusions in Onuf's nucleus in motor neuron disease (amyotrophic lateral sclerosis).

This study describes an ultrastructural examination focused on motor neurons in Onuf's nucleus in the spinal cord of four control patients without neurological disease (45-70 years) and six motor neuron disease (MND) patients (38-79 years; duration 8 months-19 years) who showed no somato-vesical dysfunction. Prompted by recent studies suggesting some sphincteric motor neurons may succumb to MND, this study sought to determine whether the wider population of neurons in Onuf's nucleus display ultrastructural cytopathology which is normally undetectable in histological preparations. Spinal cords were removed 3-20 h after death, and 1 mm slices of cord rapidly fixed in modified Karnovsky medium were processed for both light- and electronmicroscopy. 'Control motor neurons' had intact neuronal and nuclear membranes. Nissl bodies chiefly comprised ordered structures of alternate lamellae of rough endoplasmic reticulum and arrays of polyribosomes. The Golgi complexes consisted of multilamellated curvilinear stacks of ER. No intraneuronal filamentous or Bunina body inclusions were observed, but occasional axonal spheroids were seen in the neuropil. In MND, histological evidence of sparing in Onuf's nucleus was associated with abnormal ultrastructure of the motor neurons. Some sphincteric neurons were atrophic, whereas in the others, Nissl bodies were reduced in number, showed loss of structural organization or comprised polyribosomal aggregates. Golgi complexes had disrupted lamellated organization or consisted solely of distended ER. Intraneuronal filamentous Lewy-body or skein-like inclusions and Bunina bodies were identified in Onuf's nucleus of three subjects (duration of MND 8 months-2 years). The results of the present study indicate that Onuf's nucleus is vulnerable in MND, and preservation of sphincter function with qualitative histological evidence of 'sparing' does not necessarily imply a corresponding lack of ultrastructural cytopathology in this nucleus.

Adult↗
PubMed1994

Neurofilament reorganisation and neurofilament antigen redistribution in spinal motoneurones following retrograde axonal transport of diphtheria toxin.

Single unilateral injections of diphtheria toxin (DTX) into the external anal sphincter muscle or internal intercostal nerve of cat induced characteristic ultrastructural lesions in corresponding ipsilateral spinal motoneurones 6-8 days later. The chief neuronal lesion was a progressive disruption of Nissl body composition and organisation, which between days 8-19 post injection was accompanied by a progressive accumulation of neurofilaments in motoneuronal perikarya and dendrites. Some axons in the ipsilateral ventral horn became hypertrophied due to neurofilamentous accumulation. Related immunocytochemical investigations 6-35 days after injection of DTX revealed abnormal immunoreactivity intoxicated motoneurones for 200-kDa and 160-kDa phosphorylated neurofilament proteins, but not in contralateral motoneurones. By day 35 abnormal neurofilament immunostaining also occurred in ipsilateral and some contralateral interneurones but not contralateral motoneurones. Abnormalities of Nissl body endoplasmic reticulum, neurofilament organisation, and neurofilament protein immunostaining were identical after either intraneural and intramuscular injections of DTX, indicating abnormalities were attributable to toxicity and not injection-related axonal damage. Since DTX acts specifically in the soma to inhibit protein synthesis, neurofilament abnormalities are secondary to cytotoxicity and probably result from deficits in transference of existing partially phosphorylated neurofilaments to the axonal transport system, or axonal transport per se.

Animals↗
PubMed1992

Presynaptic terminal loss from alpha-motoneurones following the retrograde axonal transport of diphtheria toxin.

Intercostal motoneurones intoxicated following intraneural injection of diphtheria toxin exhibited a progressive dilatation and fragmentation of Nissl body rough endoplasmic reticulum (rER), coupled with two different forms of presynaptic terminal response. Firstly, terminal dysjunction without prior degeneration, and secondly, Wallerian-type degeneration. Dysjunction was attributed to a toxin-related failure by the motoneurones to maintain postsynaptic site structure. Degeneration was considered to arise from toxicity in presynaptic neurones, either neighbouring motoneurones or local interneurones. Morphometry revealed that by 8 days, intoxicated motoneurones exhibited a 33% loss in terminal frequency, a 15% loss in residual presynaptic membrane, and a 43% loss in overall presynaptic input. The concomitant loss of synaptic sites was greater that the overall loss of presynaptic membrane, indicating a toxin-related deficiency of the maintenance of postsynaptic sites. Analyses of the relationship between changes in terminal numbers and the development of Nissl body abnormality in the postsynaptic motoneurone identified three groups of motoneurones: (i) those with normal presynaptic input and normal neuronal Nissl body rER; (ii) those showing a dramatic loss of presynaptic input and a marked dilatation and fragmentation of Nissl bodies; and (iii) neurones exhibiting a maintained or further loss of presynaptic input coupled with extreme dilatation and fragmentation of Nissl body rER with loss of Nissl body structure. These changes are discussed in context with the known molecular action of diphtheria toxin.

Animals↗
PubMed1992

Ultrastructure of pre-synaptic input to motor neurons in Onuf's nucleus: controls and motor neuron disease.

Ultrastructural analyses of sphincteric motoneurons in Onuf's nucleus at S2 were undertaken in human spinal cord obtained 3-6 h post-mortem from three subjects with no neurological disease ('controls') and five in which death was due to motor neuron disease (MND). Neurons in specified locations within Onuf's nucleus of control subjects ranged between 17.8 and 71.7 microns diameter (mean 38.6 microns). Analyses of synaptology revealed five ultrastructural classes of presynaptic terminal synapsing with the neuronal surface membrane. When classified by size, vesicle morphology, and synaptic site structure these conformed to the S, F, T, M and C-terminals defining somatic motoneurons. No terminals characteristic of autonomic motoneurons were found. In MND subjects, neurons in Onuf's nucleus at S2 were preserved despite a paucity of neurons in medial and lateral motor nuclei and were of similar size range to those in control subjects. The morphological classes of pre-synaptic terminal found in controls, also characterized sphincteric motoneurons in MND subjects, including the C-type terminal. The presence of C-terminals indicates (i) that sphincteric motoneurons are somatic alpha-motoneurons, and (ii) that hypotheses explaining the survival of sphincteric motoneurons in MND on the basis of Onuf's nucleus being an extension of the pre-ganglionic parasympathetic nucleus, or having intrinsic autonomic properties are incorrect.

Adult↗
PubMed1991

An in vitro brainstem preparation preserving peripheral auditory function.

We describe an in vitro preparation of the brainstem and auditory periphery of the turtle. Properties of cochlear nucleus units recorded from this preparation are consistent with studies of both behavioural responses and auditory nerve fibre activity. A modified preparation for in vitro HRP tracing is also described.

Acoustic Stimulation↗
PubMed1990

Morphometric evidence from C-synapses for phased Nissl body response in alpha-motoneurones retrogradely intoxicated with diphtheria toxin.

Diphtheria toxin (DTX) kills cells by inactivating ribosomal translocation and when used to retrogradely intoxicate cat intercostal motoneurones produces marked morphological alterations in Nissl bodies, including those specifically sited postsynaptic to C-type axon terminals. Here, qualitative examinations of 'intoxicated' postsynaptic Nissl bodies reveal a progressive structural alteration marked by rER dilatation, rER lamellae fragmentation but retention of both the highly ordered multilamellate organization and ribosomal attachment until final stages of Nissl body dissolution. Morphometric results identified 3 broad phases to the postintoxication response which differed in the degree of rER cisternal dilation, and the numerical and spatial relationships between rER-lamellae, rER-bound ribosomes and rER-associated polyribosomes. These phases reflect the known molecular basis of diphtheritic toxicity and contrast with the fast developing Nissl body reaction associated with the neurotoxin ricin which also invokes ribosomal dysfunction and has been used to mimic certain features of motor neurone disease. The cytopathology of DTX and ricin are compared in the Discussion.

Animals↗
PubMed1988

Quantitative synaptology of feline motoneurones to external anal sphincter muscle.

Motoneurones innervating the cat external anal sphincter muscle were labelled retrogradely following intramuscular injections with horseradish peroxidase (HRP). Labelled motoneurones were examined by correlative light and electron microscopy (LM and EM) with special regard to a qualitative and morphometric analysis of the axon terminals resident on the neuronal membrane. By LM, labelled motoneurones were (1) ipsilateral to the injections; (2) all in S1-S2; (3) found only in the superior dorsomedial region of Onuf's nucleus; and (4) exhibited a broad spectrum of diameters (25-72 micron, mean 47.4 +/- 11.3 micron). By EM, axon terminals on the neuronal membrane when classified according to size, vesicle shape, and synaptic complex ultrastructure conformed to the S-, F-, T-, M-, and C-type terminals previously described for cat lumbosacral motoneurones. C-terminals confirmed these sphincteric motoneurones to be skeletomotor. Pooled data from midnuclear sections through 15 random labelled motoneurones (20-64-micron diameter) revealed that S- and F-type terminals predominated, with numerically few M and C types. Notwithstanding their low frequency (0.3/100 micron membrane) C-terminals contributed 1% of the mean areal coverage by terminals, which implies a potentially larger synaptic influence relative to other terminal types. Linear relationships occurred between terminal frequency (or cover) and motoneurone diameter. While motoneurones greater than 40 micron in diameter exhibited all five terminal types, labelled motoneurones less than or equal to 30 micron generally possessed only S-, F-, and occasional T-type terminals, and in this respect resembled gamma motoneurones.

Anal Canal↗
PubMed1988

Feline 'C'-type terminals possess synaptic sites associated with a hypolemmal cistern and Nissl body.

Previous findings suggest that axosomatic 'C'-type terminals on rat spinal motoneurones possess 'active' synaptic sites associated with the characteristic subsynaptic cistern and postsynaptic Nissl body, but 'C'-terminals in cat do not. A re-examination of feline 'C'-terminals undertook to compare the synaptic ultrastructure in aldehyde-fixed material stained by either osmium or ethanolic phosphotungstic acid (E-PTA). In agreement with previous findings osmicated cat 'C'-terminals failed to reveal synaptic complexes in regions possessing the subsynaptic cistern ('cisternal regions'). In contrast, cisternal regions of E-PTA stained 'C'-terminals exhibited a linear array of presynaptic dense projection all opposed by a single extended postsynaptic density which abutted directly onto both postsynaptic membrane and the cistern. This close topographical relationship is suggested to be functionally significant in the trans-synaptic trophism previously demonstrated for feline 'C'-terminals.

Animals↗
PubMed1985

Target dependence of Nissl body ultrastructure in cat thoracic motoneurones.

The effects on Nissl body (NB) ultrastructure of muscle reinnervation or neuroma formation were determined in cytochemically identified cat thoracic motoneurones subjected to axotomy by either nerve crush or nerve section with proximal ligation. Normal NB ultrastructure comprised highly ordered lamellae of rough endoplasmic reticulum (RER) with associated linear arrays of unbound polyribosomes. This ultrastructural orderliness was lost following axotomy, with or without light microscopic chromatolysis. While NBs were seen in the light microscope at late stages following both nerve crush or ligation, normal NB ultrastructure was only observed following nerve crush. An inductive effect of the periphery on NB ultrastructure is proposed and the implication of NB ultrastructure discussed in relation to protein synthesis.

Animals↗
PubMed1984

A structured program in BASIC for cell morphometry: its application to the spinal motoneurone.

The BASIC program developed for use on an RML380Z computer collects, sorts, corrects and analyzes counts of organelle number, and measurements of membrane length and profile area. These fundamental parameters are used to derive intermediate parameters and their interrelationships are examined. Data input routines permit either measurements obtained from a digitizer tablet or the use of non-automated methods. A main program controls overall program flow and discrete sub-programs implement data input, data checking, derivation of parameters and analyses. The program is applied here to analyses of motoneurone synaptology, but with simple adaptation may be used to analyze other cell types.

Animals↗
PubMed1983

Trophism between C-type axon terminals and thoracic motoneurones in the cat.

1. Quantitative ultrastructural examinations of axon terminals synapsing with normal alpha-motoneurones in segment T9 of cat spinal cord provided estimates of their numbers, sizes and synaptic structure. One synapse, the C type, derived from short-axon propriospinal segmental interneurones, was studied in detail.2. The numbers, sizes and post-synaptic structure of normal C-type synapses at T9 were compared with similar estimates from material provided by cats subjected to partial central deafferentation by double spinal hemisection at T5 and T10 between 7 days and 2 years previously.3. The proportion of C-type synapses present progessively increased from 1% in normal cats to 8.8% 200 days following hemisection, and had still attained a level of 3.1% by 2 years; these increases imply that the absolute number of C-type synapses underwent increase.4. Mean sizes of C-type synapses increased from 4.0 mum (normal) to 5.8 mum (200 days) and retained their enlarged sizes up to 2 years (5.9 mum). Furthermore, while 84% of C-type synapses were under 6 mum in length in normal motoneurones, 48% were over 6 mum long 200 days post-operatively.5. The unique post-synaptic structure of C-type synapses also proliferated following partial central deafferentation of the motoneurones. The elongated cistern, increased numbers and individual lengths of lamellae of the associated underlying rough endoplasmic reticulum indicated a trophic interaction between the presynaptic C terminal and its post-synaptic motoneurone.6. Counts of ribosomes ;bound' to lamellae of the subsynaptic rough endoplasmic reticulum, and of the lamellae-associated polyribosomes interposed between individual lamellae for normal and 200 day post-operative C-type synapses indicated an over-all post-operative increase in capacity for local subsynaptic protein synthesis topographically directed towards this type of axon terminal.7. The observed greater increase in frequency of ribosomes ;bound' to the rough endoplasmic reticulum, together with an over-all proliferation of this structure, specificially indicated an increased capacity for synthesis of protein for utilization in sites remote from those of synthesis (e.g. a trans-synaptic passage of protein).8. A hypothesis is advanced on the basis of the above results relating both pre- and post-synaptic changes in structure to an increased functional activation of the segmental short-axon propriospinal interneurones forming the C-type synapses, as a compensatory response to partial central deafferentation of spinal motoneurones.

Afferent Pathways↗
PubMed1982

A structured program in BASIC for the analysis of peripheral nerve morphometry.

The BASIC program developed for use on a M.S.I. 6800 microprocessor analyses cross-sectional measurements of fibre (D) and axon (d) calibre made on electron micrographs of suitable magnification. Data input routines cover measurements of area and diameter, made manually or presented as fully computed values printed out by a M.O.P. Separate sub-routines input digitized data from a digitizer tablet coupled to the microprocessor. Myelin sheath thickness (2M) (D-d), or area and 'g' (d/D) are derived. The two analytical programs provide either frequency distributions, with associated statistics for each parameter, or analyses of the relationships (D:d), (d:2M) and (D:g). The highly structured nature of the program permits it to be edited or extended according to the project. The main programme controls the overall flow of analysis; discrete subprograms inserted into the main program perform the data input, parameter derivations, data checks and analyses. This system offers maximum flexibility and is suitable for most morphometric investigations involving peripheral nerves.

Animals↗
PubMed1981

A quantitative ultrastructural study of dorsal root regeneration.

Regeneration in rat lumbo-sacral dorsal roots was studied 5-71 days following crush lesions. Wallerian degeneration occurred up to 20 days. At 11 days degenerating myelin was found in both Schwann cells and macrophages. Myelination was first observed 4 mm central to the crush at 7 days, and myelin became compact when the mesaxon exhibited 3.5 turns about the axon (about 11 days post-operatively). At 71 days, 69% of all fibres were myelinated, compared with 36% in normal roots. An example of 2 axons myelinating within the same Schwann cell occurred at 20 days. In normal roots curvilinear relationships were found between axon diameter and fibre diameter, myelin thickness and axon diameter, and between g and fibre diameter. In contrast, linear relationships between these parameters occurred in post-operative roots up to 71 days. Curvilinearity returned at 71 days. Alterations in the relationship between axon diameter and myelin thickness during regeneration indicated that myelin growth lagged behind axon growth throughout, but was more noticeable in larger calibre fibres. By 71 days, larger fibres exhibited disproportionately thin myelin, whilst small fibres possessed abnormally thick myelin compared to normal fibres of similar calibre. Regeneration was limited by axons failing to make successful central synaptic connections and by the poor metabolic response of dorsal root ganglion cells to sectioning of their central processes.

Animals↗
PubMed1977

The distribution and relative sized of fibre types in the extensor digitorum longus and soleus muscles of the adult rat.

Histochemical fibre types classified in sections stained for succinic dehydrogenase (sdh) and myosin ATPase at pH 9.4, were found to be distributed in a consistent manner within the extensor digitorum longus (EDL) and 4 soleus muscles of the adult rat. Simple morphometric techniques applied to complete transverse sections of both muscles showed that the relative distributions and proportions of fibre types along their deep to superficial, and medial to lateral, axes varied accoording to the histochemical method used for fibre typing. Similar differences occurred when the relative ranges of size exhibited by each fibre type were compared in sections stained for SDH and ATPase, and the discrepancies in fibre classification were confirmed by an analysis of individual fibres in serial sections. The findings are discussed in relation to those previosly reported for the EDL and soleus muscles of the rat.

Adenosine Triphosphatases↗
PubMed1977

The distribution and relative sizes of three histochemical fibre types in the rat tibialis anterior muscle.

A quantitative study of the tibialis anterior muscle of the adult rat showed that the proportions of each 'histochemical' fibre type varied between adjacent regions along the deep to superficial and medial to lateral axes of the muscle. The distribution of fibre types classified with oxidative enzymes was similar to that using phosphorylase, but differed from that obtained with sections stained for myosin ATPase. This apparent discrepancy in classifation was confirmed by comparison of the ranges of fibre cross sectional areas for each fibre type classified on the basis of their oxidative enzme and ATPase activites, and by an independent analysis of individual fibres in serial sections stained for succinic dehydrogenase and ATPase.

Adenosine Triphosphatases↗