Cyto- and Dendroarchitecture

The laminar distribution of spinal neurons has been widely accepted. Its main advantage is its simple and comprehensive scheme of spinal cord organization and physiological properties can also be correlated to this structural arrangement.

Cytoarchitectural laminae are characterized by the density and topography of spinal neurons in the grey matter and can usually be identified on thick cross sections (fig. 1). In addition, each lamina has its own characteristics which are particularly distinct at the level of cervical and lumbar enlargements. Most of the information about dendritic territories has been obtained by using Golgi impregnation methods. In addition to the laminar arrangement in the coronal plain, in the ventral horn the cervical and lumbar motoneurons form rostrocaudal motor columns⁷ (fig. 1).

Figure 1

a) Cross section of the lumbar portion of the rat spinal cord showing the layered arrangement of the hemicord. The white matter is separated from the grey matter by a broken line. LSN= lateral spinal nucleus which takes place outside the grey matter in (more...)

Lamina I is the dorsalmost lamina which covers the tip of the dorsal horn. It has a loosely packed neuropil and a low neuronal density with neurons of variable size and distribution. The most typical neuron is the so-called Waldeyer⁸ cell: large, fusiform neuron with disk-shaped dendritic domain.^9-12 However, in cat and rat also small and medium-sized pyramidal neurons were identified in this lamina^{3,10,11,13,14} and characterised as fusiform, pyramidal and multipolar cells.¹³

Lamina II appears as a darkly stained band in Nissl-stained sections due to its high neuronal density (substantia gelatinosa, Rolando, 1824). In cat and rodents the inner and outer zones can be distinguished^2,3 although in humans there is not a clear separation between these zones. The neuronal population consists of small fusiform neurons. There are two main cell types which form the majority of the population of lamina II: the islet cells with a rostrocaudal axis and the stalked cells with a dorsoventral dendritic tree. Other types of neurons have been described such as arboreal, curly, border, vertical, filamentous and stellate cells.^9,15-18 It is possible, however, that some of these latter neurons correspond to each other or to the two main cell types. Islet cells contain GABA therefore they are considered as the inhibitory cells of this lamina.

Lamina III can be easily distinguished from lamina II by its lower neuronal density and by the presence of intermediate size neurons. This layer has a mixed population of antenna-like and radial neurons.^19-21 These cells have a simpler dendritic morphology than those in layer II.^9,22,23 Many of the above cells contain inhibitory neurotransmitters: GABA or glycine.²⁴

Lamina IV in man and cat has a variety of antenna-like cells and the so-called transverse cell.^7,9,23 Most of their dendrites originate dorsally on the cell body and spread towards lamina II and III. In animals, the axons of lamina IV neurons mainly enter the spinocervical tract, which is vestigial in humans. Most probably human lamina IV neurons project to the spinothalamic tract.²⁵ Laterally from this lamina there is a small group of neurons embedded in the lateral funiculus: the lateral spinal nucleus²⁶ (fig. 1). Its neurons project to the midbrain and brainstem and send processes to lamina IV itself.

Lamina V-VI have a similar cyto- and dendroarchitecture. The medial part contains fusiform and triangular neurons. The lateral part is not clearly separated from the dorsolateral funiculus. This part corresponds to the reticular formation in the brainstem and consists of medium-sized multipolar neurons.

Lamina VII occupies the intermediate zone of the grey matter and is formed by an homogeneous population of medium-sized multipolar neurons. In the appropriate segments it contains some well-defined nuclei, such as the intermediolateral nucleus (T1-L1; medially) and the dorsal nucleus of Clarke (T1-L2; laterally). The intermediolateral nucleus plays a role in the autonomic sensory and motor functions and the axons of neurons from the dorsal nucleus of Clarke form the ascending fibres of the dorsal spinocerebellar tract.

Lamina VIII has, unlike laminae I-VII a dorsoventral extension. It contains a variety of neurons with dorsoventrally polarized dendritic tree. The largest multipolar neurons can be distinguished from motoneurons only by their finer Nissl bodies by using conventional morphological techniques.²⁷

Lamina IX is made up of groups of cells that form motor nuclei. Motoneurons have a unique position in this lamina, being the only spinal cord neuron which has its axon almost entirely in the peripheral nervous system. The α-motoneurons have the largest somata in the cord (50 x 70 μm) whilst the γ-motoneurons are smaller. Motoneurons can be easily recognized by the abundance of Nissl bodies in their cytoplasm and their multipolar shape (fig. 1). Their dendrites extend for long distances, dorsally as far as lamina VI. Small neurons at the medial border of the motor nucleus are identified as the short-axoned inhibitory interneurons, the Renshaw cells. Although Rexed's classification^2,3 did not differentiate between motoneuron groups in lamina IX, these neurons can be divided into four separate columns in the human cord: the ventromedial, ventrolateral, dorsolateral and central columns²⁸ (fig. 2).

Figure 2

Schematic representation of the dendroarchitecture of spinal motoneurons in various motor columns. The ventromedial motoneurons (IX-vm) form vertical and longitudinal dendritic branches (not shown), motoneurons in the ventrolateral (IX-vl) and central (more...)

Motoneurons projecting to the axial muscles are found in the ventromedial column,^28,29 those innervating proximal musculature of the limbs occupy medial and ventral position while neurons innervating distal limb muscles are located in dorsal and lateral positions. In all but one (dorsolateral) motoneuron column the dendritic polarization is longitudinal and dendritic trees overlap for a long distance (fig. 2). Such a dendritic organization favours synchronization and synergy for axial, proximal and calf muscles.^4,5 In contrast to these columns, motoneurons in the dorsolateral column have radially oriented dendritic trees without much overlap of their dendrites (fig.2). This dendritic arrangement favours precise contacts with segmental afferents and may contribute to a more precise control of movements of distal muscles.

Lamina X

This lamina corresponds to the substantia grisea centralis, the grey matter around the central canal. Two cell types can be recognized: (1) Bipolar cells with fan-shaped dendritic tree (dorsal portion of lamina X) and (2) bipolar cells with poorly ramified longitudinal dendrites (ventral portion).³⁰

Interneurons in the Spinal Cord

Interneurons are probably the most important modulating cell types in the spinal cord. The importance of spinal interneuronal networks has only recently been acknowledged although the flexibility of these networks became apparent as early as in the 1950s.

Initially only electrophysiological approaches were used, later the precise location, morphology and immunohistochemical features helped to distinguish special interneuronal classes.

The very first morphologically and physiologically identified interneurons were the Renshaw cells and Ia interneurons (Renshaw cells project on motoneurons and thus establish the recurrent inhibition, whereas Ia interneurons are activated by Ia afferents of agonist muscles and inhibit antagonistic motoneurons).^31,32 Renshaw cells, Ia and Ib inhibitory interneurons, interneurons in disynaptic and polysynaptic reflex pathways and interneurons mediating descending commands were the “classical interneurons” and their function was thoroughly analyzed in a series of studies. Recently a number of new interneurons modulating special functions were described, e.g.,interneurons involved in the clasp-knife reflex, bladder function, control of respiration and last-order premotor interneurons, etc. It is expected that the number of these highly specialized interneurons will further increase with time (for recent reviews and references see Jankowska 2001,³³ Edgley 2001³⁴).

Most interneuron types have also been characterised by their neurochemical features. Renshaw cells, for example, express not only glycine, their characteristic inhibitory neurotransmitter, but they reportedly synthesize calcium binding proteins calbindin-D28k and parvalbumin.^35-37

This short description of spinal interneurons suggests that the fine control of spinal functions mostly depends on the integrity of spinal interneuronal networks. It should be noted that interneurons named after their characteristic input (Ia, Renshaw, etc) receive a variety of multisensory inputs of different origins and these inputs together determine what the interneuron actually will do.

Intrinsic Pathways

These tracts not only establish connections between different neuronal groups and segments of the spinal cord but also act as relays between descending pathways and intrinsic spinal neurons. Accordingly, well defined ascending and descending white matter bundles are committed to propriospinal functions.

The Lissauer's tract tract can be localized between the entering dorsal roots and lamina I. It is mainly composed of unmyelinated descending and ascending fibres and both types extend a few segments. The majority of these fibres originate from the dorsal roots whilst the rest is intrinsic in nature terminating on marginal and substantia gelatinosa cells. The comma tract is a comma-shaped thin fibre bundle in between the fasciculi cuneatus and gracilis. It contains descending fibres from the cervical dorsal roots. The septomarginal tract is situated in the dorsal white matter and its position varies at the level of different segments. It consists of descending dorsal root and intrinsic fibres. The cornucommissural tract can be found along the dorsal commissure and contains ipsilaterally running descending and ascending propriospinal fibres. The anterior and lateral ground bundles are present throughout the spinal cord being most developed at the levels of enlargements. They contain both ascending and descending long and short fibres. They originate in the ipsilateral hemicord and terminate throughout the grey matter.

Ascending Pathways

The ascending pathways are formed by the central axons of dorsal root ganglion cells entering the spinal cord via the dorsal roots. They either enter an ascending fibre tract (dorsal column pathways) or terminate in the spinal grey matter. About two-third of these fibres are fine, unmyelinated, slowly-conducting C fibres. The myelinated fibre components can be classified as fast-conducting, large, myelinated Aβ, and slower-conducting, thinly myelinated Aδ fibres. Primary sensory fibres either terminate in the dorsal column nuclei of the medulla or in the superficial dorsal horn according to a segregated pattern. Thin fibres related to temperature and pain terminate in laminae I and II, whereas coarse fibres terminate in deeper layers (laminae III-V) and in the ventral horn as well (proprioceptive afferents). Furthermore, primary afferents coming from cutaneous receptors terminate almost exclusively in lamina II in rat and cat whilst visceral and muscle afferent terminals are mainly confined to laminae I and V.^42,43

The dorsal column pathways include the medially located fasciculus gracilis (Goll) and the laterally situated fasciculus cuneatus (Burdach). The fasciculus gracilis contains dorsal root afferents from the lower limbs and lower part of the body, the fasciculus cuneatus from the upper limb and upper part of the trunk. The fibres synapse on neurones of the nucleus gracilis and nucleus cuneatus, respectively. These pathways play role in discriminative sensory tasks, such as two-point discrimination, detection of speed and direction of movements and judging of cutaneous pressure.⁴⁴ The spinothalamic tract originates from neurons in laminae I,V,VII and VIII,²⁵ however the distribution of spinothalamic neurons shows significant species differences. In humans the axons cross to the ventrolateral column and terminate in the ventral posterolateral and in the central lateral nuclei of the thalamus. In other mammals they terminate mainly in the posterior thalamic nuclear complex. Functionally, this tract conveys the accurate localization of pain and thermal stimuli. Ventrolateral cordotomies presented evidence that other tracts may also transmit pain stimuli.⁴⁵ The spinoreticular tract originates from cells situated bilaterally throughout the spinal grey matter. The ascending fibres in the ventral and lateral funiculi terminate in several nuclei of the reticular formation. Many spinothalamic ascending fibres also give collaterals to reticular nuclei. This pathway is responsible for carrying a variety of sensory information. The spinocervicothalamic tract uses an intermediate nucleus in the spinal cord, the lateral cervical nucleus, which is consistent in lower mammals but often absent in human spinal cords. Afferent fibres to this nucleus arise from the ipsilateral lamina IV in all cord segments. Neurons from the lateral cervical nucleus project to the contralateral thalamus via the medial lemniscus. This system is involved in tactile conditioned reflexes, tactile and proprioceptive placing and size discrimination. The spinocerebellar tracts (dorsal and ventral) carry information primarily arising from the lower extremities. The dorsal spinocerebellar tract is formed by axons of the ipsilateral nucleus dorsalis of Clarke (present in Th₁-L₂ segments in humans) and projects to the vermis and the paravermal regions of the cerebellum. It conveys information from muscle spindles, Golgi tendon organs, joints and mechanoreceptors of the lower extremities. Axons of cells situated in laminae V and VII in the lumbosacral spinal cord form the ventral spinocerebellar tract. It projects to the vermis and paravermal region of the cerebellum and probably carries information about the interrelationship of different muscle groups. Equivalent information from the upper extremities are conveyed by the cuneocerebellar and the rostral spinocerebellar tracts of the spinal cord.

Descending Pathways

The corticospinal tract is most developed in higher primates and species differences are most pronounced for this tract. The cells of origin are located in the motor cortex and their axons form the pyramidal tract. In most mammals fibres from neurones in the postcentral gyrus also contribute to this tract. In humans the bulk of the fibres cross in the lower medulla and form the lateral corticospinal tract whereas uncrossed fibres remain in the ventral funiculus and then cross in the ventral commissure. In some species the organization of this tract is different.^46,47 Functionally, the corticospinal pathway exerts a fine and amplified motor control by influencing other descending pathways.^48,49 Fibres of the reticulospinal tracts originate from the dorsal and central parts of the medulla and the pontine tegmentum. The terminal distribution of medial reticulospinal fibres is very dense in the ventral horn of the enlargements while the lateral reticulospinal tract fibres terminate in laminae I and V.⁵⁰ Fibres of the vestibulospinal tract originate from the lateral and medial vestibular nuclei. Both lateral and medial tract fibres terminate ipsilaterally in laminae VII and VII and form mono- or polysynaptic inhibitory connections with motoneurons, especially with those of neck and back muscles. The rubrospinal tract is well developed in lower mammals and less developed in humans. Its fibres originate from the caudal magnocellular part of the red nucleus and project according to a somatotopic pattern contralaterally to laminae V-VII. In cat there is a direct rubrospinal connection to motoneurons.⁵¹ The tract exerts excitatory effects on flexor motoneurons and inhibits extensor motoneurons. The tectospinal tract tract originates from the superior colliculus and terminates contralaterally in the ventral horn of the upper cervical cord where its fibres establish multisynaptic connections with motoneurons of neck muscles.

Apart from the major descending tract, there are many minor fibre bundles originating from the interstitial nucleus of Cajal, solitary and retroambiguous nuclei, and the paraventricular nucleus of the hypothalamus. Noradrenergic fibres descend from the locus coeruleus and the lateral pontine nuclei to the grey matter and to the intermediolateral nucleus, respectively. Serotonergic projections arise from the raphe magnus and raphe pallidus and obscurus nuclei terminate either in laminae I and V (raphe magnus fibres) or in the ventral horn (rest of the fibres).

Pain and Temperature

The peripheral receptors for various modalities of sensation are specialised sense organs that are contacted by axons from dorsal root ganglion neurons. These neurons have a peripheral process and a central branch that enters the spinal cord where they branch. These neurons that are directly linked with the peripheral structures are called first order neurons, and their role in processing of sensory information is largely determined by their branching pattern. Figure 4a illustrates some of the sense organs of the first order neurons that are involved in pain and temperature sensation and also shows that the main target of the branches of the central portion of this first order neuron terminates and synapses on neurons in the substantia gelatinosa. It is from this part of the dorsal horn where the second order neurons give rise to their processes which convey the information to other parts of the spinal cord and brain. However, there are ascending and descending branches of the second order neurons that synapse on cells in different segments of the spinal cord and on more ventral interneurons that are concerned with control of movement and integration of somatic afferent inputs with those from other parts of the central nervous system.⁵² Thus these second order neurons play a crucial role in the processing of sensory information within the spinal cord. Not only somatic afferent fibres converge into the neurons in the substantia gelatinosa, but visceral sensation and pain also converges onto this group of second order neurons. In addition there is a strong input from various structures of the brain that impinge upon neurons in the substantia gelatinosa modify the input from the periphery and in this way the outcome of sensation (for further reading see Brown 1991,⁵³ Schomburg 1990⁵⁴). It is partly because of this convergence of inputs to this part of the spinal cord that sensation is not simply the result of particular peripheral inputs.

Figure 4

a) and b) illustrates schematically the types of sensory nerve endings in peripheral tissues innervated by sensory nerves (peripheral processes). It also shows the central processes and their lamination in the spinal cord and medulla. The insert on the (more...)

Touch and Tactile Discrimination

The sensation of light touch is initiated from specialized sense organs in the skin or connective tissue or from free nerve endings in the dermis. The sense organs are contacted by axons from the cells of dorsal root ganglia and the information reaches the spinal cord via the central branch of the neurons of the dorsal root ganglion cells. These central branches form long tracts which give off branches to interneurons of the posterior horn in laminae VI and VII. The second order neurons within the spinal cord that process information about touch are thus in lamina VI and VII.

The same structures that are involved in the sensation of touch are also contributing to more sophisticated sensory functions such as two point discrimination, awareness of movement of body parts, as well as the position of various body parts in relation to each other. However these functions are also critically dependent on proprioception.

Proprioception

The sense organs that convey this modality of sensation are located in muscles, tendons and joints (Figs. 4b and 5). The structure of these is rather complex and indicate their important function in conveying the initial signal. In the muscle the annulospiral and flower spray endings of the spindles are monitoring muscle length and this task is complicated by the fact that the spindles are themselves a group of muscle fibres ensheathed in a connective tissue capsule and contacted by 2 types of sensory fibres. In addition to their sensory innervation the muscle fibres within the spindle receive their own motor innervation from small motoneurons and axons referred to as gamma efferents. Thus by relaxing or contracting the muscle fibres within the spindle the message about the state of the muscle is modified even before it reaches the spinal cord. In muscle tendons there are organs (Golgi tendon organs) which monitor the stretch imposed upon the tendon, and the Pacinian corpuscules within joints and close to bony structures monitor the pressure exerted upon these structures. The axons of sensory nerves that carry the information from the spindles towards the spinal cord are among the largest and fastest conducting nerves in the body. The central branch make up the medial division of the dorsal root as it enters the spinal cord. The central branch splits after entering the spinal cord and some of these enter the anterior horn where they synapse directly onto motoneurons to initiate a monosynaptic reflex, or onto interneurons to exert via interneurons more sophisticated control over locomotor activity.⁵⁵ These monosynaptic connections are rather unique in that there is a high degree of specificity and muscle spindle afferents from a given muscle contact only motoneurons that innervate the muscle of the origin of this afferent input. Other branches enter the posterior funiculus and ascend towards Clarke's nucleus in the posterior grey horn. Some descending and ascending branches synapse on interneurons in laminae V, VI, and VII. Axons of these cells cross the midline and ascend in the ventral spinocerebellar tract to communicate with the cerebello-olivary system.

Figure 5

a) and b) The sensory and motor innervation of the mammalian muscle spindle is illustrated in a. This figure also shows the schematic picture of the cross section of the spinal cord with the various inputs to γ-motoneurons (left) and α-motoneurons (more...)

Thus the various parts of the sensory system inform the brain about the external and internal stimuli impinging onto the extremities and trunk. However this information undergoes considerable processing by the circuitry of the spinal cord and is continuously modified by it.⁵⁴

Reflexes

Our understanding of spinal cord physiology has until recently been dominated by observations of Sherrington⁵⁶ (1910) and his colleagues that the structures of the spinal cord are able to produce stereotyped responses to external stimuli. These responses were referred to as reflexes and carefully defined and observed. The simplest of these reflexes is the monosynaptic stretch reflex, elicited by activation of the IA afferent fibres that originate from the muscle spindle, and when activated produces contraction of the synonymous muscle. However even the study of this simple reflex revealed a great degree of complexity in the spinal cord circuitry. The strength of muscle contraction in response to the same stimulus was not always the same and was influenced by preceding activity of the spinal cord. In order to explain some of the findings associated with the variability of reflex activity it was necessary to consider events such as temporal and spatial summation of excitatory inputs, and inhibitory influences from other sources.Thus even the simplest “reflex” turned out to display considerable variability.⁵⁶ Nevertheless the information about the behaviour of the structures that mediate the responses to various stimuli in the spinal cord obtained by the study of reflex activity was of immense importance. It taught us that the observation of temporal and spatial summation of excitatory inputs is caused by the ability of neurons to add up excitatory postsynaptic potentials (EPSPs) and therefore when 2 inputs, each of which is too weak to produce a response on its own, impinge upon a neuron simultaneously, or with a slight delay, they can produce a response since the depolarisation of the cell reaches a threshold level which fires off an action potential. These rules apply even in the case of the simplest reflex response such as the stretch reflex, which is monosynaptic and the integration is carried out by only one cell, the motoneuron. All other reflexes are polysynaptic, and therefore each neuron involved in the response can contribute to the final outcome i.e., the motor response to a particular stimulus (see fig. 5). The study of these relatively simple spinal reflexes revealed other features of the system, i.e., that neurons are not only excited, but can be inhibited by particular inputs. Such inhibition is either postsynaptic so that the membrane potential of the postsynaptic neuron increases and thus the same excitatory input fails to depolarize the neuron sufficiently to initiate an action potential, or inhibition can be presynaptic, by which the amount of excitatory transmitter released from the presynaptic terminal is reduced.

Patterned Movements Organised by the Circuitry of the Spinal Cord

In addition to the monosynaptic stretch reflex the circuitry of the spinal cord can generate patterned responses that involve movement of several joints. The best explored reflex of this type is the flexor, or withdrawal reflex in response to various sensory stimuli, and in particular in response to pain. During this reflex the extremity is withdrawn from the site of the stimulus. The flexor reflex is a complex movement which involves a highly organized sequence of activation and inhibition of motoneurons to particular muscles. It affects muscles of the contralateral limb so that the animal is supported during the time when the limb is involved in the flexor reflex and is lifted off the ground. Another patterned response that can be organized by the spinal cord is stepping. In acutely spinalised animals Brown⁵⁷ (1911) showed that the spinal cord could trigger rhythmic walking movements. These movements are of interest, since they do not depend entirely on sensory inputs and are generated by neurons located in the spinal cord. The group of neurons responsible for the organisation of this movement has been referred to as central pattern generator (CPG).⁵⁸

Most of the information on spinal cord CPGs in mammals has been obtained on experimental animals such as rats or cats. However, whether the spinal cord of primates and humans is able to produce the same responses when disconnected from the brain is less well documented. So far the available information suggests that the isolated spinal cord of primates or humans is unable to generate such primitive stepping movements as those described for the cat.⁵⁹ Nevertheless some spinal reflex responses are preserved after complete spinal cord lesion in humans. These include the stretch reflex, which is often exaggerated and the flexor reflex. However, these responses are not stereotyped and change when they are elicited repetitively.⁶⁰ Thus even the human spinal cord is able to generate complex responses, which are influenced by repeated activity, by mechanisms that we do not understand.

The localisation of supraspinal locomotor regions is well established in the sense that electrical stimulation of such regions can elicit walking, or even galloping in decorticate cats suspended above a treadmill belt.⁶¹ Stimulation of these areas in primates prepared in a similar manner as cats, also elicited walking and trotting. However the monkeys walked on all 4 limbs.⁶² Thus like in cats the mesencephalic locomotor centre was able to activate the locomotor function of the primate spinal cord, but without the connection with this centre the stimuli that induced locomotor activity in the spinal cat were unable to do so in the spinal monkey.

Until now this section described the potential of the spinal cord to produce integrated responses without depending on the influences from the brain. However it is important to emphasize that this situation is rare and even after spinal cord injury in man the separation of the spinal cord from the brain is rarely complete. It is therefore important to consider spinal cord function in relation to the control systems that normally regulate its performance. Figure 6 summarizes the various influences from the higher centres that may influence the performance of the spinal cord circuitry.

Figure 6

The scheme illustrates the various structures involved in the control of locomotor function.

Since this book is concerned with the possibility that neuronal or glial transplants will either replace damaged parts of the spinal cord, or encourage existing structure to regenerate or resume their function it seems pertinent to mention observations that concern the importance of various descending pathways for recovery of locomotor activity. It appears that in patients with spinal cord injuries the preservation of the ventral funiculi is best correlated with recovery of gait,⁶³ while patients with well preserved sensation of touch and position, but severe damage to the anterior part of the cord have a poor chance to regain the ability to walk.⁵⁹ In monkeys trained to walk on a treadmill return of locomotor performance after spinal cord injury was critically dependent on the preservation of at least one ventrolateral funiculus. Retrograde labelling of the preserved funiculus showed that the axons in the preserved funiculus originates in the vestibular, reticular and raphe nuclei.⁶² Thus it appears that these structures are of critical importance for the control of the spinal cord central pattern generator. In the context of the topic discussed in this book this finding is of utmost importance, for it indicates that for successful restoration of motor function efforts should be made to reconnect particular structures, rather then haphazardly investigate indiscriminate growth of axons that may not be able to contribute to improvement of function.