Mark B. Plenderleith

The plant lectin Bandeiraea simplicifolia I-B4 identifies a subpopulation of small diameter primary sensory neurones which innervate the skin in the rat

In recent years a variety of glycoconjugates have been found associated with the plasma membrane of mammalian primary sensory neurones. The functional significance of these glycoconjugates remains obscure but their carbohydrate chains have been widely implicated in cell-cell recognition (or adhesion) during development. The plant lectin Bandeiraea simplicifolia I-B4 identifies a characteristic galactose-containing glycoconjugate associated with small diameter primary sensory neurones. In this study we have used a combination of lectin binding and retrograde tracing to examine the distribution of this glycoconjugate on sensory neurones which innervate different target tissues in the rat. The fluorescent tracer diamidino yellow was applied the cut end of peripheral nerves which selectively innervate either the skin, muscle or viscera. Retrogradely labelled neurones were then screened for lectin binding using a lectin-horseradish peroxidase conjugate. The results revealed that Bandeiraea simplicifolia I-B4 binding is associated with over one-third of neurones which innervate the skin, but only a small proportion of neurones innervating muscle or viscera.

Binding sites for the plant lectin Bandeiraea simplicifolia I-isolectin B4 are expressed by nociceptive primary sensory neurones

Circumstantial evidence suggests that binding sites for the plant lectin Bandeiraea simplicifolia I-isolectin B(4) are expressed by nociceptive primary sensory neurones. In order to test this hypothesis directly, we have used a combination of intracellular staining of functionally characterised primary sensory neurones and lectin binding. Consistent with the hypothesis, none of the low threshold primary sensory neurones we sampled expressed lectin binding sites, whilst a subpopulation of the nociceptive neurones did.

Ultrastructural analysis of the central terminals of primary sensory neurones labelled by transganglionic transport of bandeiraea simplicifolia I-isolectin B4.

In this study the ultrastructural appearance of primary sensory neurones labelled by the injection of the plant lectin Bandeiraea simplicifolia I-isolectin B(4) (BSI-B(4)) into a peripheral nerve has been examined in the rat. Electron microscopy of the somata of retrogradely labelled neurones showed the lectin to be associated with the inner surface of cytoplasmic vesicles, supporting the premise that the uptake of BSI-B(4) into sensory neurones is by the process of receptor-mediated endocytosis. Light and electron microscopic analysis of the spinal cord revealed transganglionically transported lectin in unmyelinated axons in the dorsolateral funiculus and axon terminals concentrated mainly within lamina II of the dorsal horn. Detailed analysis of 1377 of these axon terminals revealed that the majority were glomerular in shape and surrounded by up to 14 other unlabelled profiles. These findings suggest that primary sensory neurones which transganglionically transport BSI-B(4) have a synaptic ultrastructure similar to that which has been previously reported for unmyelinated primary sensory neurones. Moreover, it appears that the axon terminals of these neurones are subjected to extensive modulation. Examination of the vesicle content of lectin labelled axon terminals revealed that the majority contained small agranular vesicles while large granular vesicles were observed only occasionally. These findings support the suggestion that the populations of neurones expressing binding sites for BSI-B(4) are fairly distinct from those containing neuroactive peptides. In conclusion, the results of the current study suggest that the lectin BSI-B(4) can be used as a histological marker for a subpopulation of small diameter primary sensory neurones and provide further evidence for the potential of this lectin as a useful tool in the study of pain.

Analysis of the distribution of binding sites for the plant lectin Bandeiraea simplicifolia I‐isolectin B4 on primary sensory neurones in seven mammalian species

The purpose of the present study was to investigate the binding patterns of the plant lectin Bandeiraea simplicifolia I‐isolectin B4 (BSI‐B4) to sensory neurones in seven mammalian species. The dorsal root ganglia and spinal cords of three rats, mice, guinea pigs, rabbits, flying foxes, cats, and marmoset monkeys were screened for BSI‐B4 using lectin histochemistry. BSI‐B4 binding was associated with the soma of predominantly small‐diameter primary sensory neurones in the dorsal root ganglia and their axon terminals within laminae I and II of the superficial dorsal horn in all seven species. The similarities of lectin binding patterns in each of these species suggest that the glycoconjugate to which BSI‐B4 binds has a ubiquitous distribution in mammals, and supports the proposal that this lectin may preferentially bind to a subpopulation of sensory neurones with a similar functional role in each of these species. Anat Rec 268:105–114, 2002.

Expression of lectin binding in the superficial dorsal horn of the rat spinal cord during pre- and postnatal development

The plant lectin Bandeiraea simplicifolia I-B4 binds to the soma and central terminals of a subpopulation of unmyelinated primary sensory neurones in the adult rat. The binding site of this lectin is thought to be the terminal alpha-D-galactose residue of a membrane associated glycoconjugate which may be involved in the development of specific connections between small diameter primary sensory neurones and second order neurones in the superficial dorsal horn of the spinal cord. To begin to investigate this possibility we have examined the development of lectin binding in the dorsal horn of pre- and postnatal rats. Lectin binding first appeared on axon profiles in the superficial dorsal horn of the spinal cord at embryonic days 18/19. Previous studies in the rat have revealed that the central processes of small diameter primary sensory neurones enter the dorsal horn at embryonic days 18/19. Our findings suggest that the glycoconjugate to which this lectin binds, is expressed by the central processes of small diameter primary sensory neurones as they grow into the spinal cord. It is therefore possible that this glycoconjugate is involved in the development of topographically ordered neural connections within the dorsal horn of the spinal cord.

Histochemical localisation of a galactose‐containing glycoconjugate expressed by sensory neurones innervating different peripheral tissues in the rat

Abstract  The plant lectin Bandeiraea simplicifolia I‐isolectin B4 (BSI‐B4) identifies a galactose‐containing, membrane‐associated glycoconjugate expressed by a discrete subpopulation of unmyelinated primary sensory neurones in the rat. We have previously suggested that BSI‐B4 selectively binds to primary sensory neurones that innervate the skin. However, in that study, the tracer diamidino yellow was applied to the cut ends of peripheral nerves to identify neurones innervating particular target tissues. In this study, we have avoided axotomy by retrogradely labelling primary sensory neurones from peripheral tissues using the carbocyanine dye 1,1′‐dioctadecyl‐3,3,3′,3′‐tetramethylindocarbacyanine perchlorate (DiI). DiI was injected into the plantar skin, gastrocnemius muscle, and pyloric region of the stomach in rats. Corresponding ganglia were sectioned, incubated in BSI‐B4 conjugated to fluorescein isothiocyanate, and examined with a fluorescence microscope. DiI‐labelled cells were identified by red fluorescence within the cytoplasm, whereas cells binding BSI‐B4 displayed green fluorescence associated with the plasma membrane and Golgi apparatus. Quantitative analysis revealed that 36.2% of cutaneous neurones, 7.6% of muscle neurones, and 6.8% of visceral neurones expressed the BSI‐B4‐binding site, indicating that a small but significant proportion of small‐diameter primary sensory neurones innervating muscle and viscera also express BSI‐B4‐binding sites.

Induction of partial immunity in both males and females is sufficient to protect females against sexual transmission of Chlamydia

Sexually transmitted Chlamydia trachomatis causes infertility, and because almost 90% of infections are asymptomatic, a vaccine is required for its eradication. Mathematical modeling studies have indicated that a vaccine eliciting partial protection (non-sterilizing) may prevent Chlamydia infection transmission, if administered to both sexes before an infection. However, reducing chlamydial inoculum transmitted by males and increasing infection resistance in females through vaccination to elicit sterilizing immunity has yet to be investigated experimentally. Here we show that a partially protective vaccine (chlamydial major outer membrane protein (MOMP) and ISCOMATRIX (IMX) provided sterilizing immunity against sexual transmission between immunized mice. Immunizing male or female mice before an infection reduced chlamydial burden and disease development, but did not prevent infection. However, infection and inflammatory disease responsible for infertility were absent in 100% of immunized female mice challenged intravaginally with ejaculate collected from infected immunized males. In contrast to the sterilizing immunity generated following recovery from a previous chlamydial infection, protective immunity conferred by MOMP/IMX occurred independent of resident memory T cells. Our results demonstrate that vaccination of males or females can further protect the opposing sex, whereas vaccination of both sexes can synergize to elicit sterilizing immunity against Chlamydia sexual transmission.

Analysis of the unmyelinated primary sensory neurone projection through the dorsal columns of the rat spinal cord using transganglionic transport of the plant lectin Bandeiraea simplicifolia I-isolectin B4

We have examined the projection of unmyelinated primary sensory neurones through the dorsal columns of the rat spinal cord using transganglionic transport of the plant lectin Bandeiraea simplicifolia I-isolectin B4. A small volume of the lectin was injected into the sciatic nerve of anaesthetised rats to label the central terminals of nociceptive primary sensory neurones. Following a survival period of 7 days, transverse and longitudinal sections of the superficial dorsal horn, dorsolateral funiculus and the dorsal columns from spinal segments L4 through to T13 were screened for lectin transport using light and electron microscopy. Longitudinal sections of the thoraco-lumbar region of spinal cord were also examined for lectin binding. Light and electron microscopy revealed transganglionically transported and bound lectin in the superficial dorsal horn and dorsolateral funiculus of the L3 and L4 segments of spinal cord. However, no lectin transport or binding was observed within the dorsal columns at any level of spinal cord examined. From these results, we suggest that the unmyelinated neurones within the dorsal columns do not express the binding site for BSI-B4 and, as such, may be responsible for visceral rather than cutaneous sensation. In line with the theories regarding a postsynaptic dorsal column pathway, these results suggest that nociceptors that bind BSI-B4 are not involved in a direct ascending projection through the dorsal columns.

Distribution of Binding Sites for the Plant Lectin Ulex europaeus Agglutinin I on Primary Sensory Neurones in Seven Different Mammalian Species

There is an increasing body of evidence to suggest that different functional classes of neurones express characteristic cell-surface carbohydrates. Previous studies have shown that the plant lectin Ulex europaeus agglutinin-I (UEA) binds to a population of small to medium diameter primary sensory neurones in rabbits and humans. This suggests that a fucose-containing glycoconjugate may be expressed by nociceptive primary sensory neurones. In order to determine the extent to which this glycoconjugate is expressed by other species, in the current study, we have examined the distribution of UEA-binding sites on primary sensory neurones in seven different mammals. Binding sites for UEA were associated with the plasma membrane and cytoplasmic granules of small to medium dorsal root ganglion cells and their axon terminals in laminae I–III of the grey matter of the spinal cord, in the rabbit, cat and marmoset monkey. However, no binding was observed in either the dorsal root ganglia or spinal cord in the mouse, rat, guinea pig or flying fox. These results indicate an inter-species variation in the expression of cell-surface glycoconjugates on mammalian primary sensory neurones.

Use of Lectins as Transganglionic Neuronal Tracers in the Study of Unmyelinated Primary Sensory Neurons

One of the major advances in neurobiology in the last two decades has been the development of neural tracing techniques that allow the investigator to unequivocally establish the precise pattern of connections between different populations of neurons. Early neuronal tracers were relatively nonspecific agents transported by all neurons at the injection site. More recently however, the affinity of plant lectins for the membrane-associated glycoconjugates expressed by neurons has been utilized to develop a class of neuronal tracers that are taken up and transported by specific populations of neurons. The effectiveness of plant lectins as neuronal tracers is dependent on the distribution of the membrane-associated glycoconjugates to which they bind If the glycoconjugate to which the lectin binds is present on the majority of neurons, then this lectin will be suitable for use throughout the nervous system. However if the lectin binds to a relatively discrete group of neurons, it may be used to examine the connections of this specific population of neurons.