Elsevier

Progress in Neurobiology

Volume 54, Issue 5, 5 March 1998, Pages 531-548
Progress in Neurobiology

The problem of assessing effective neuroprotection in experimental cerebral ischemia

https://doi.org/10.1016/S0301-0082(97)00078-6Get rights and content

Abstract

In animal models of global and focal ischemia neuroprotection is typically determined by quantifying the degree of cell loss or reduction in infarct volume shortly after the ischemic insult. These methods are unable to reliably detect more subtle forms of neuronal death and dysfunction that arise from injury to non-homogenous cell populations (e.g. hilar and striatal neurons), or to dendrites (e.g. loss of structural proteins or decreased synaptic transmission). It is argued that this type of covert injury contributes to a wide range of functional impairments (e.g. decreased working memory, altered field potentials, loss of forelimb dexterity) that are rarely used as outcome measures in experimental studies even though they are of paramount importance clinically.

The limitations of a purely histological approach in assessing neuroprotection are clearly illustrated using examples of protective drug therapies, mild hypothermia and ischemic preconditioning. An alternative strategy that incorporates behavioural, electrophysiological and histological endpoints is put forth as a more powerful method for gauging neuroprotection. The strength of this approach will be increased if these assessments are performed on the same animals. By incorporating functional measures and longer postischemic survival into their experimental protocols, investigators will increase the validity of their models and hopefully reduce the likelihood of advancing ineffective therapies into costly clinical trials.

Introduction

Stroke or cerebral ischemia is a leading cause of death and permanent disability for which there is currently no effective treatment. New drug therapies and other interventions have emerged that may limit this type of brain injury (Koroshetz and Moskowitz, 1996). However, these therapies are typically evaluated in animal stroke models simply by counting the remaining number of healthy cells or calculating infarct volumes 1–7 days after the ischemic episode. Recent research suggests that some treatments may only delay cell death, whereas others leave cells in a functionally abnormal state. In this review we examine the pitfalls associated with current methods of gauging neuroprotective efficacy in both global and focal models of ischemia. An alternative evaluative approach is put forth that incorporates behavioural, neurophysiological and histological indices of neuroprotection.

Section snippets

The issue of postponed cell death

Histological endpoints (i.e. cell counts) determined up to 7 days after ischemia are customarily used to assess the efficacy of potential therapeutic interventions. A major problem with this strategy is that a number of drugs and other treatments appear to only postpone cell death (Morse and Davis, 1990; Li and Buchan, 1995; Nurse and Corbett, 1996; Dietrich et al., 1993; Colbourne and Corbett, 1994; Valtysson et al., 1994; Corbett and Crooks, 1997). Thus, cells that appear preserved by some

Histological assessment

The first concern with histological assessment is: what to count and what to measure? It is well known that CA1 neurons are highly susceptible to global ischemic injury and their laminar distribution and large size make them relatively easy to quantify. However, other neuronal populations (e.g. CA2 and hilar neurons) are equally, if not more sensitive, to global ischemia (Hsu and Buzsaki, 1993; Matsuyama et al., 1993; Akai and Yanagihara, 1993). Yet neuronal loss within these regions is rarely

Neuropathology

In general, the pattern of neuronal injury in gerbil and rat models of global ischemia is very similar (Pulsinelli et al., 1982; Kirino and Sano, 1984; Smith et al., 1984; Crain et al., 1988) with CA1, CA2, hilar, dorsolateral caudate and cortical neurons (in middle layers) being the most vulnerable. Due to its incomplete circle of Willis, it is easy to produce near total forebrain ischemia in the gerbil by occluding the carotid arteries for either 3 or 5 min. The ensuing loss of CA1 neurons,

Neuropathology

Most models of focal ischemia involve permanent or temporary occlusion of the MCA (Tamura et al., 1981; Buchan et al., 1992; Longa et al., 1989) that supplies the lateral surface of the cerebral cortex and a limited number of subcortical structures, most notably the caudate nucleus (Scremin, 1995). These cortical and striatal areas are known to be involved in the processing and integration of sensory-motor information (Whishaw et al., 1986; Andersen et al., 1991) and thus most investigators

Electrophysiology

The previous sections discussed the importance of using behavioural tests in conjunction with a histological evaluation following global and focal ischemia. However, in most behavioural tests there is often considerable recovery following ischemia. In many cases, deficits in behavioural tasks are only observed within the first few weeks after the insult (Markgraf et al., 1992; Andersen et al., 1991; Corbett et al., 1992). Recent studies have provided evidence that neuronal death can continue

Conclusions

It should be clear from the foregoing discussion of global and focal ischemic studies that assessing neuroprotection solely on the basis of conventional histological procedures (e.g. Nissl stains) is not adequate. Examples were presented where different treatments (hypothermia, ischemic preconditioning and neuroprotective drugs) appeared to convey neuroprotection when histological endpoints were used but considerably less efficacy was found with functional endpoints. To some extent these

Acknowledgements

This work was supported by grants from the Medical Research Council of Canada, the Heart and Stroke Foundation of Newfoundland and Labrador and the Astra Canadian Neuroscience Network awarded to DC. The authors would like to thank Jennifer Dowden and Paul Dooley for helpful comments on this manuscript.

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