Animal model of posterior cingulate cortex hypometabolism implicated in amnestic MCI and AD

Abstract

The posterior cingulate cortex (PCC) is the brain region displaying the earliest sign of energy hypometabolism in patients with amnestic mild cognitive impairment (MCI) who develop Alzheimer’s disease (AD). In particular, the activity of the mitochondrial respiratory enzyme cytochrome oxidase (C.O.) is selectively inhibited within the PCC in AD. The present study is the first experimental analysis designed to model in animals the localized cortical C.O. inhibition found as the earliest metabolic sign of early-stage AD in human neuroimaging studies. Rats were used to model local inhibition of C.O. by direct injection of the C.O. inhibitor sodium azide into the PCC. Learning and memory were examined in a spatial holeboard task and brains were analyzed using quantitative histochemical, morphological and biochemical techniques. Behavioral results showed that sodium azide-treated rats were impaired in their memory of the baited pattern in probe trials as compared to their training scores before treatment, without non-specific behavioral differences. Brain analyses showed that C.O. inhibition was specific to the PCC, and sodium azide increased lipid peroxidation, gliosis and neuron loss, and lead to a network functional disconnection between the PCC and interconnected hippocampal regions. It was concluded that impaired memory by local C.O. inhibition in the PCC may serve to model in animals a metabolic lesion similar to that found in patients with amnestic MCI and early-stage AD. This model may be useful as an in vivo testing platform to investigate neuroprotective strategies to prevent or reduce the amnestic effects produced by posterior cingulate energy hypometabolism.

Introduction

Minoshima et al., 1994, Minoshima et al., 1997 were the first to report that metabolic energy reduction in the posterior cingulate cortex (PCC) is the first sign of brain hypometabolism very early in Alzheimer’s disease (AD). Neuroimaging evidence for an early metabolic lesion in PCC has also been found in asymptomatic subjects at genetic risk for AD, such as subjects homozygotic for the E4 allele of the apolipoprotein E gene (Reiman et al., 1996, Small et al., 2000), and in patients with amnestic mild cognitive impairment (MCI) who later develop AD (Mosconi, 2005). Moreover, recent diffusion tensor imaging showed that the cingulum fibers that connect the PCC with hippocampal regions have reduced integrity in both MCI and AD patients (Zhang et al., 2007). This is consistent with a disruption in the functional correlations of a network of brain regions involving the PCC and interconnected medial temporal regions in MCI patients at risk for AD (Sorg et al., 2007).

Human studies support a role of the PCC in spatial orientation and memory, which are affected in amnestic MCI and early-stage AD. Indeed, PET and MRI studies showed that the severity of AD symptoms is correlated with hypometabolism in PCC but not temporal regions (Alsop et al., 2000, Hirono et al., 1998, Ishii et al., 1997). Cammalleri et al. (1996) described a patient with a tumor in the PCC who was unable to navigate in its surroundings due to the inability to use landmarks for route finding. Functional studies in healthy subjects also provide evidence for a role of the PCC in visually-guided behavior relating to aspects of navigation and memory. A spatial attention task showed fMRI activation in the PCC that was strongly correlated with the speed of target detection when spatial cues were present (Mesulam, Nobre, Kim, Parrish, & Gitelman, 2001). The PCC was also active when subjects were asked to remember visual landmarks and their movements along a previously learned route (Berthoz, 1997). Similarly, Maguire (1997) found that the PCC was involved during tasks that required learning a room that contained salient objects and empty rooms only distinguished by their shape, as well as remembering environments recently learned and those very familiar. In particular, PET studies showed that the PCC is critically involved in a network for memory retrieval (Nyberg et al., 1996, Cabeza et al., 1997).

Animal studies also suggest a role of the PCC in spatial navigation and memory. Single-cell recordings show that specific sensory stimuli can activate cingulate neurons that are sensitive to the direction the animal is facing (Chen, Lin, Green, Barnes, & McNaughton, 1994). Furthermore, the PCC may specifically be involved during the late stages of learning. In an active avoidance task, discriminative neuronal activity (cells firing to a stimulus that predicts shock) occurs late in learning in the PCC (Gabriel, 1993). This is supported by studies showing that lesioning this area before task acquisition impaired rats only during the late stages of acquisition (Bussey et al., 1996, Bussey et al., 1997). Lesioning the PCC with NMDA impaired rats’ performance in the radial arm maze, water maze, and an object-in-place task, whereby objects are switched in location in the second trial (Vann & Aggleton, 2002). Aspiration of the PCC produced deficits in the water maze and matching-to-place tasks (Harker and Whishaw, 2002, Sutherland et al., 1988, Whishaw et al., 2001).

The evidence from human and animal studies indicates that the PCC is involved in spatial navigation, perhaps with a bigger contribution during the stages of memory retrieval. Human functional neuroimaging shows that this area is the earliest affected metabolically in patients with AD (Minoshima et al., 1997) and in patients with MCI who later develop AD (Mosconi, 2005). Importantly, Valla, Berndt, and Gonzalez-Lima (2001) found that the underlying biochemical defect for PCC hypometabolism in AD brains was a selective activity reduction of the mitochondrial enzyme cytochrome oxidase (C.O.). This respiratory enzyme is responsible for the activation of oxygen for aerobic energy metabolism, and it provides an index of neuronal metabolic energy capacity because it is essential for ATP production in mitochondria (Wong-Riley, Nie, Hevner, & Liu, 1998). The energy hypometabolism in the PCC of AD patients is specifically linked to decrement in C.O. activity, it is not found in age-matched control brains, and it is not secondary to decrement in other mitochondrial enzymes or the number of amyloid plaques or neurofibrillary tangles in PCC (Valla et al., 2001). Furthermore, C.O. activity in the superficial layers of PCC in AD brains showed a progressive reduction tightly linked to disease duration, which was not observed in other cortical regions such as the motor cortex (Valla et al., 2001).

There is a need to model in animals PCC hypometabolism similar to that linked to amnestic MCI and early-stage AD to be able to investigate neuroprotective strategies to prevent or reduce the amnestic effects produced by PCC hypometabolism. The goal of this study was to determine whether an amnestic mild impairment could be induced in rats by partial inhibition of C.O. activity in the PCC, and whether this local metabolic lesion would lead to network functional dissociation between the PCC and interconnected hippocampal regions implicated in the progression of AD (Sorg et al., 2007, Zhang et al., 2007). PCC hypometabolism was produced by sodium azide, a mitochondrial toxin that selectively inhibits C.O. activity by blocking the transfer of electrons to oxygen (Bennett et al., 1992, Cada et al., 1995). The objective was to model the specific metabolic lesion linked to early-stage AD by inhibiting C.O. activity in the PCC, not just damage this region by other means in a non-specific manner. This study tested this hypothesis by locally inhibiting C.O. activity in the PCC after rats were trained in a spatial food search task and examining its effects on memory retrieval and regional brain metabolism.