Richard J. Riopelle

Survival of Cholinergic Forebrain Neurons in Developing p75NGFR-Deficient Mice

The functions of the low-affinity p75 nerve growth factor receptor (p75NGFR) in the central nervous system were explored in vivo. In normal mice, approximately 25 percent of the cholinergic basal forebrain neurons did not express TrkA and died between postnatal day 6 and 15. This loss did not occur in p75NGFR-deficient mice or in normal mice systemically injected with a p75NGFR-inhibiting peptide. Control, but not p75NGFR-deficient, mice also had fewer cholinergic striatal interneurons. Apparently, p75NGFR mediates apoptosis of these developing neurons in the absence of TrkA, and modulation of p75NGFR can promote neuronal survival. Cholinergic basal forebrain neurons are involved in learning and memory.

Deficits in saccadic eye-movement control in Parkinson's disease

In contrast to their slowed limb movements, individuals with Parkinsons disease (PD) produce rapid automatic eye movements to sensory stimuli and show an impaired ability to generate voluntary eye movements in cognitive tasks. Eighteen PD patients and 18 matched control volunteers were instructed to look either toward (pro-saccade) or away from (anti-saccade) a peripheral stimulus as soon as it appeared (immediate, gap and overlap conditions) or after a variable delay; or, they made sequential saccades to remembered targets after a variable delay. We found that PD patients made more express saccades (correct saccades in the latency range of 90-140 ms) in the immediate pro-saccade task, more direction errors (automatic pro-saccades) in the immediate anti-saccade task, and were less able to inhibit saccades during the delay period in all delay tasks. PD patients also made more directional and end-point errors in the memory-guided sequential task. Their inability to plan eye movements to remembered target locations suggests that PD patients have a deficit in spatial working memory which, along with their deficit in automatic saccade suppression, is consistent with a disorder of the prefrontal-basal ganglia circuit. Impairment of this pathway may release the automatic saccade system from top-down inhibition and produce deficits in volitional saccade control. Parallel findings across various motor, cognitive and oculomotor tasks suggest a common mechanism underlying a general deficit in automatic response suppression.

Depressive Symptoms in Parkinson's Disease: A Comparison With Disabled Control Subjects

A high incidence of depressive symptoms has been observed in patients with Parkinsons disease (PD). PD involves a loss of central monoamines, and a decrease of monoamines has been implicated in depression; therefore, it is possible that depressive symptoms in PD result from the loss of endogenous neurotransmitters. However, it is equally possible that depressive symptoms represent a reaction to the chronic disabling course of PD. By comparing depressive symptoms in PD patients to those in matched patients with other chronic disabling diseases not involving a loss of central monoamines, it may be possible to decide between these alternatives. Thus, depressive symptoms were assessed in 45 patients with PD and 24 disabled controls that did not differ from the PD subjects on a measure of functional disability. Results showed that PD subjects obtained significantly higher total scores on the Beck Depression Inventory (BDI) than controls. PD subjects scored significantly higher than controls on BDI items grouped to reflect cognitive-affective and somatic depressive symptoms. The BDI scores of PD subjects were not reliably related to age, sex, duration of PD, or clinical ratings of PD symptom severity or functional disability. Self-rated disability and the number of recent medical problems were the greatest predictors of depressive symptoms. These findings supported the hypothesis that depressive symptoms in PD may not represent solely a reaction to disability. (J Geriatr Psychiatry Neurol 1989; 2:3-9).

Developmental regulation of nerve growth factor and its receptor in the rat caudate-putamen

In prior studies, nerve growth factor (NGF) administration induced a robust, selective increase in the neurochemical differentiation of caudate-putamen cholinergic neurons. In this study, expression of NGF and its receptor was examined to determine whether endogenous NGF might serve as a neurotrophic factor for these neurons. The temporal pattern of NGF gene expression and the levels of NGF mRNA and protein were distinct from those found in other brain regions. NGF and high-affinity NGF binding were present during cholinergic neurochemical differentiation and persisted into adult-hood. An increase in NGF binding during the third postnatal week was correlated with increasing choline acetyltransferase activity. The data are consistent with a role for endogenous NGF in the development and, possibly, the maintenance of caudate-putamen cholinergic neurons.

Axotomy-Induced Apoptotic Cell Death of Neonatal Rat Facial Motoneurons: Time Course Analysis and Relation to NADPH-Diaphorase Activity

Rapid and massive death of motoneurons occurs following axotomy in neonatal mammals. This likely results from the neurons being deprived of access to target-derived trophic factors, as their death can be prevented by application of a variety of neurotrophic factors to the proximal end of the cut nerve. Since trophic factor-deprived embryonic chick motoneurons undergo apoptosis in vitro, we have investigated whether axotomized neonatal rat facial motoneurons undergo apoptotic cell death in vivo. Following facial nerve transection during the first postnatal day, the dying motoneurons show characteristic morphological changes of apoptosis and undergo DNA fragmentation, as detected by an in situ end labeling technique. An initial sharp burst of DNA fragmentation, between 12 and 24 h postaxotomy, accompanies a steep decline in neuronal numbers, indicating that neuronal cell death rapidly follows endonuclease cleavage of DNA. However, the interval between axotomy and onset of DNA fragmentation varies widely. By 4 days postnatum only 38% of the lesioned motoneurons have survived an initial rapid phase of neuronal loss, whereas 11% survive to 10 days postnatum at least. NADPH-diaphorase/nitric oxide synthase activity has been implicated as having a causal role in the death of lesioned motoneurons. We have found that there is a sustained increase in the intensity of NADPH-diaphorase histochemical staining in axotomized neonatal facial motoneurons, but that this is first detectable well after the onset of DNA fragmentation and cell death. This suggests that nitric oxide, or its metabolites, does not initiate cell death in this model.

RECIPROCAL MODULATION OF TRKA AND P75NTR AFFINITY STATES IS MEDIATED BY DIRECT RECEPTOR INTERACTIONS

Equilibrium binding of 125I‐nerve growth factor (125I‐NGF) to cells coexpressing the tyrosine kinase receptor A (TrkA) and common neurotrophin receptor (p75NTR), cells coexpressing both receptors where p75NTR is occupied, and cells expressing only p75NTR, revealed reciprocal modulation of receptor affinity states. Analysis of receptor affinity states in PC12 cells, PC12 cells in the presence of brain‐derived neurotrophic factor (BDNF), and PC12nnr5 cells suggested that liganded and unliganded p75NTR induce a higher affinity state within TrkA, while TrkA induces a lower affinity state within p75NTR. These data are consistent with receptor allosterism, and prompted a search for TrkA/p75NTR complexes in the absence of NGF. Chemical crosslinking studies revealed high molecular weight receptor complexes that specifically bound 125I‐NGF, and were immunoprecipitated by antibodies to both receptors. The heteroreceptor complex of TrkA and p75NTR alters conformation and/or dissociates in the presence of NGF, as indicated by the ability of low concentrations of NGF to prevent heteroreceptor crosslinking. These data suggest a new model of receptor interaction, whereby structural changes within a heteroreceptor complex are induced by ligand binding.

Adhesion properties of a neuronal epitope recognized by the monoclonal antibody HNK-1

A carbohydrate epitope on adhesion proteins of the developing nervous system, and on myelin-associated glycoprotein, is recognized by the monoclonal antibody HNK-1. The HNK-1 epitope bearing proteins and the monoclonal antibody alter, in a dose-dependent manner, the interaction between neurons and neurite-promoting substrate-attached materials released from cultured neural cells.

Functional interactions of neuronal heparan sulphate proteoglycans with laminin

Quantitative biosynthetic studies using cellular extracts and neuron conditioned medium demonstrated that heparan sulphate proteoglycans (HSPGs) comprised 20-25% of the sulphated proteoglycans produced by neurons while the remainder consisted of chondroitin sulphate proteoglycans (CSPGs). When chromatographic fractions containing guanidine extracted and partially purified proteoglycans from culture medium conditioned by neurons (NCM) were used to pretreat a laminin substrate, neurite formation by sensory neurons was enhanced. Enhanced neurite promoting activity was not apparent if, during the pretreatment of the laminin substrate with NCM, heparan sulphate glycosaminoglycans (HS) were present. To determine the molecular basis of cell surface HSPG interactions with immobilized laminin, adhesion and neurite growth by dissociated sensory neurons were quantified at 4 h in vitro--a time at which there was no apparent contribution of released proteoglycans to neurite growth. Whereas adhesion was not influenced, neurite growth was partially inhibited in a dose-dependent manner if the sensory neurons were coincubated with HS, and if the cells were pretreated, prior to seeding, with heparitinase. The inhibitory effect produced by coincubation with saturating concentrations of HS was no longer apparent if the cells had been pretreated with heparitinase. These findings distinguish quantitatively between neurite growth on laminin and on laminin-HSPG complexes, and suggest that some neuronal cell surface and released HSPGs are involved in neurite growth by virtue of non-covalent interactions with glycosaminoglycan binding domains of laminin.

Peripheral nerve contains heterogeneous growth factors that support sensory neurons in vitro.

The availability of dissociated cultures of embryonic neurons has given impetus to a number of studies which have detected trophic factors in many mammalian and avian tissues maintained in vivo and in vitro [1, 2, 4-8, 10, 11, 13, 16, 18-21]. Using a single sensory neuron biological assay for mouse nerve growth factor (NGF) [17] as a reference standard, we report here that neurite promoting activity is highly enriched in segments of adult mouse peripheral nerve. Neurite outgrowth at 30 h, kinetics of neurite appearance and inhibition with rabbit antiserum to mouse NGF indicate that the trophic activities are heterogeneous and support both NGF-dependent and NGF-independent neuron populations of chick embryo sensory ganglia.

MOLECULAR CORRELATES OF SPINAL CORD REPAIR IN THE EMBRYONIC CHICK : HEPARAN SULFATE AND CHONDROITIN SULFATE PROTEOGLYCANS

Proteoglycan (PG) biosynthesis in vivo and PG-associated neurite growth-promoting activity in vitro were examined in the thoracic spinal cord of embryonic chick at times during which functional recovery following axonal damage is permitted, and at later times when such functional recovery is restricted. Over a 10-day period encompassing the permissive and restrictive periods the ratio of newly synthesized heparan sulfate (HS) PG to chondroitin sulfate (CS) PG decreased by more than 50%. Specific PG-associated neurite-promoting activity (NPA) of a PG fraction immobilized on a laminin substrate was 75-fold higher at E9 than at E17. Perturbations of the two families of PGs indicated that all laminin-bound NPA was associated with HSPGs from E9 cord, and that removal of the influence of CSPGs from PG extracts of E17 cord unmasked neurite-promoting activity on a poly-D-lysine substrate of the same magnitude as that observed on a laminin substrate. Neurite-promoting activity associated with HSPGs and high HS to CS ratios of newly synthesized PGs characterize the permissive period for axonal regeneration in the chick embryo spinal cord. In the restrictive period for axonal regeneration neurite promoting activity is masked by the presence of CSPGs which are synthesized at higher levels than HSPGs.