Karen M. Doyle

Unfolded proteins and endoplasmic reticulum stress in neurodegenerative disorders

•  Introduction •  ER stress and the UPR ‐  The IRE1 axis: non‐conventional splicing of XBP1 mRNA ‐  The PERK axis: attenuation of translation ‐  The ATF6 axis: regulated proteolytic activation •  ER stress–induced apoptosis •  ER stress and autophagy •  The UPR and neurodegenerative disease ‐  Alzheimers disease ‐  Parkinsons disease ‐  Amyotrophic lateral sclerosis ‐  Prion diseases •  Future perspectives

Comparison of various N-methyl-D-aspartate receptor antagonists in a model of short-term memory and on overt behaviour.

This study examined the effects on rat behaviour of antagonists acting at various sites on the N-methyl-D-aspartate (NMDA) receptor complex, i.e. the glutamate recognition site (CPP), ion channel (dizocilpine), glycine recognition site [(+)-HA-966] and the NR2B subunit-selective compound ifenprodil. Specifically, the effects of these agents were examined on working memory, assessed using the operant delayed match-to-position task (DMTP), and overt behaviour, assessed (a) in animals responding for food under a variable interval 20-s (VI20) schedule and (b) by spontaneous behaviour. Dizocilpine, CPP and (+)-HA-966 each reduced accuracy in the DMTP task independent of delay. At equivalent doses, changes in locomotor behaviour and VI20 responding were evident. In contrast, ifenprodil failed to impair accuracy in the DMTP task, even at doses that affected other performance measures and reduced VI20 responding. The relevance of these observations to neuroprotective and anticonvulsant doses of these compounds is considered.

Polyamines in the Brain: Distribution, Biological Interactions, and their Potential Therapeutic Role in Brain Ischaemia

The endogenous polyamines (spermine, spermidine, and putrescine) are present at relatively high concentrations in the mammalian brain and play crucial roles in a variety of aspects of cell functioning. Stroke is the third most common cause of death and the leading cause of disability among adults in the western world. Brain polyamine levels change dramatically following cerebral ischaemia. Polyamines may be involved in the pathophysiological processes underlying brain ischaemia through several possible mechanisms. These include direct effects on ion channels and receptors modulating potassium, and most importantly calcium trafficking, or through the production of toxic metabolites. Considerable evidence shows that the non-competitive polyamine antagonists, ifenprodil and eliprodil, are neuroprotective. Interestingly, novel polyamine analogues, such as N(1)-dansylspermine, BU36b, and BU43b, have also recently been shown to have neuroprotective potential. The exact mechanisms of the neuroprotection afforded by the polyamine antagonists and their clinical applicability is worthy of further study.

Characterisation of a functional polyamine site on rat mast cells: association with a NMDA receptor macrocomplex

Polyamines can modulate activation of N-methyl-D-aspartate (NMDA) receptors by binding to a specific polyamine site associated with a NMDA receptor macrocomplex. Polyamines induce histamine release from mast cells, although the mechanism had not been defined. We have examined whether spermine, a natural polyamine, and compound 48/80, regarded as a synthetic polyamine, activate mast cells by a polyamine site associated with a NMDA receptor macrocomplex. Spermine induced secretion of histamine from rat peritoneal mast cells and rat brain mast cells in a concentration-dependent manner. Rat peritoneal mast cells were used as a model system to explore the effects of NMDA antagonists on polyamine-induced histamine release. Ifenprodil, MK801 and arcaine inhibited histamine secretion from mast cells exposed to polyamines; the percentage inhibition was greater against spermine than compound 48/80. These data support the proposal that spermine (and possibly compound 48/80) induce histamine release from mast cells by interacting with a specific polyamine site on a NMDA receptor complex.

Investigation of the involvement of the N-methyl-D-aspartate receptor macrocomplex in the development of spermine-induced CNS excitation in vivo

1 The involvement of the N‐methyl‐D‐asparate (NMDA) receptor macrocomplex in the development of spermine‐induced CNS excitation in vivo was investigated. 2 Injection of 100 μg of spermine into the left lateral cerebral ventricle of female Laca mice (20–25 g) resulted in the development of two distinct phases of CNS excitatory effects which were quantified by a scoring system. 3 The first phase effects occurred within minutes of injection and generally lasted for about 1 h. Most mice showed scratching of the upper body, frequent face washing and some mice developed clonic convulsions. By about 2 h after injection, the second phase of effects began to develop in the form of body tremor which worsened with time and culminated in fatal tonic convulsions, generally within 8 h of injection. 4 Pretreatment of the mice with dizocilpine (0.3 mg kg−1, i.p.) resulted in antagonism of the first phase of spermine‐induced effects, but a higher dose (0.3 mg kg−1, (x2), i.p.) was necessary to inhibit the second phase effects. 5 Whereas the glutamate antagonist, 3‐((R)‐2‐carboxypiperazin‐4‐yl) propyl‐1‐phosphonic acid (D‐CPP) (10, 20 mg kg−1, i.p.), the glycine antagonist 7‐chlorokynurenate (10, 30, 50 nmol, i.c.v.), or the polyamine antagonist ifenprodil (30, 60 mg kg−1, i.p.) antagonized the first phase of effects produced by spermine, these agents given as monotherapy, were ineffective against the development of the second phase of effects. 6 Co‐administration of ifenprodil with either D‐CPP or 7‐chlorokynurenate resulted in a dose‐dependent antagonism of the development of the second phase of spermine‐induced effects. 7 It is concluded that the development of the two temporally distinct phases of spermine‐induced effects may be mediated by pharmacologically distinct mechanisms, although the results suggest that the NMDA receptor macrocomplex may be involved in both phases of effects. Furthermore, a moderate dose of D‐CPP or 7‐chlorokynurenate appears to enhance the inhibitory potential of ifenprodil in vivo.

Investigation of the actions and antagonist activity of some polyamine analogues in vivo

The ability of three putative polyamine antagonists to antagonize behavioural changes induced by spermine was assessed. Injection of an excitotoxic dose of spermine (100 μg, i.c.v.) in mice results in the development of a characteristic behavioural profile, which has two temporally distinct phases. The early events include clonic convulsions, and the later, more general excitation, includes tremor and culminates in the development of a fatal tonic convulsion. Co‐administration of arcaine (25 μg, i.c.v.) potentiated the early phase effects after spermine injection, but antagonized the development of spermine‐induced tonic convulsions. A larger dose of arcaine (50 μg, i.c.v.) given alone resulted in the development of spermine‐like body tremor and convulsions. It therefore appears that arcaine is not a pure polyamine antagonist in vivo, but may be a partial agonist. Similarly, 1,10‐diaminodecane appeared to act as a partial agonist in vivo, although it was less potent than arcaine. In contrast, diethylenetriamine (DET) effectively inhibited the development of the early effects of spermine, but was ineffective against the spermine‐induced CNS excitation and tonic convulsions. It is concluded that none of the putative polyamine antagonists tested behaved as effective polyamine antagonists in vivo, although each produced some antagonism.

Nerve growth factor-mediated inhibition of apoptosis post-caspase activation is due to removal of active caspase-3 in a lysosome-dependent manner

Nerve growth factor (NGF) is well characterised as an important pro-survival factor in neuronal cells that can inhibit apoptotic cell death upstream of mitochondrial outer membrane permeabilisation. Here we addressed the question of whether NGF can also protect against apoptosis downstream of caspase activation. NGF treatment promoted a rapid reduction in the level of the p17 subunit of active caspase-3 in PC12 cells that had been induced to undergo apoptosis by various cytotoxins. The mechanism involved TrkA-dependent activation of extracellular signal-regulated kinase (ERK1/2) but not phosphatidylinositol 3-kinase (PI3K)/Akt, and de novo protein synthesis. Involvement of inhibitor of apoptosis proteins (IAPs) and proteasomal degradation were ruled out. In contrast, inhibition of lysosome function using chloroquine and concanamycin A reversed NGF-induced removal of p17. Moreover, in NGF-treated cells, active caspases were found to be localised to lysosomes. The involvement of macroautophagy and chaperone-mediated autophagy were ruled out. Taken together, these findings suggest an anti-apoptotic mechanism by which NGF induces removal of active caspase-3 in a lysosome-dependent manner.

Fear-induced suppression of nociceptive behaviour and activation of Akt signalling in the rat periaqueductal grey: role of fatty acid amide hydrolase

The endocannabinoid system regulates nociception and aversion and mediates fear-conditioned analgesia (FCA). We investigated the effects of the fatty acid amide hydrolase (FAAH) inhibitor URB597, which inhibits the catabolism of the endocannabinoid anandamide and related N-acylethanolamines, on expression of FCA and fear and pain related behaviour per se in rats. We also examined associated alterations in the expression of the signal transduction molecule phospho-Akt in the periaqueductal grey (PAG) by immunoblotting. FCA was modelled by assessing formalin-evoked nociceptive behaviour in an arena previously paired with footshock. URB597 (0.3 mg/kg, i.p.) enhanced FCA and increased fear-related behaviour in formalin-treated rats. Conditioned fear per se in non-formalin-treated rats was associated with increased expression of phospho-Akt in the PAG. URB597 reduced the expression of fear-related behaviour in the early part of the trial, an effect that was accompanied by attenuation of the fear-induced increase in phospho-Akt expression in the PAG. Intra-plantar injection of formalin also reduced the fear-induced increase in phospho-Akt expression. These data provide evidence for a role of FAAH in FCA, fear responding in the presence or absence of nociceptive tone, and fear-evoked increases in PAG phospho-Akt expression. In addition, the results suggest that fear-evoked activation of Akt signalling in the PAG is abolished in the presence of nociceptive tone.

The pre-ischaemic neuroprotective effect of a novel polyamine antagonist, N1-dansyl-spermine in a permanent focal cerebral ischaemia model in mice

The polyamine sites on the NMDA receptor complex offer a therapeutic target for focal ischaemia, potentially devoid of most side effects associated with NMDA antagonists. In this study, we investigated the effect of a novel polyamine antagonist, N(1)-dansyl-spermine (0.5-10 mg kg(-1)) in a permanent focal cerebral ischaemia model in mice, and compared its effect to that of MK-801 (0.3-3 mg kg(-1)) following administration 30 min prior to ischaemia. A battery of histological and behavioural tests was employed following permanent middle cerebral artery occlusion to assess any neuroprotective effect. Following middle cerebral artery occlusion, N(1)-dansyl-spermine (1-5 mg kg(-1)) and MK-801 (1 or 3 mg kg(-1)) caused a comparable and significant reduction in the percentage hemisphere lesion volume. Similarly, both drugs significantly reduced oedema and neurological deficit score to a similar extent. Locomotor activity in MCAO mice was not significantly improved by MK-801 or N(1)-dansyl-spermine, although N(1)-dansyl-spermine induced a trend towards significant improvement. Significant improvement in rotarod performance was observed at neuroprotective doses with both drugs. Upon comparison of the profile of effects, N(1)-dansyl-spermine at least matched the effectiveness of MK-801 as a neuroprotective agent in this model. In addition, in sham-operated control mice, N(1)-dansyl-spermine was well tolerated, in contrast to the pronounced adverse effects of MK-801 on locomotor activity and rotarod performance. In conclusion, this study has shown that N(1)-dansyl-spermine is as effective a neuroprotective drug as MK-801 in this model. Moreover, in contrast to MK-801, N(1)-dansyl-spermine could be a promising therapeutic candidate for stroke as it is well tolerated at neuroprotective doses in sham-operated animals.

Recent advances in psychoneuroimmunology: Inflammation in psychiatric disorders

Psychiatric disorders are common and complex and their precise biological underpinnings remain elusive. Multiple epidemiological, molecular, genetic and gene expression studies suggest that immune system dysfunction may contribute to the risk for developing psychiatric disorders including schizophrenia, bipolar disorder, and major depressive disorder. However, the precise mechanisms by which inflammation-related events confer such risk are unclear. In this review, we examine the peripheral and central evidence for inflammation in psychiatric disorders and the potential molecular mechanisms implicated including inhibition of neurogenesis, apoptosis, the HPA-axis, the role of brain-derived neurotrophic factor and the interplay between the glutamatergic, dopaminergic and serotonergic neurotransmitter systems.