Review articleRegulation of object recognition and object placement by ovarian sex steroid hormones
Introduction
The object recognition (OR) task was first introduced in 1988 to provide a method of testing episodic memory in rodents that was similar to methods used in clinical neuropsychology [1]. The task capitalizes on rodent's inherent predilection for novelty. In the most common version of the task, rodents explore two identical objects during a single training session. During the single test session, subjects are then allowed to explore an object identical to the training objects and a novel object. More time spent exploring the novel object indicates memory for the familiar object. In the years since its introduction, OR has become prevalent in rodent learning and memory studies, where it is used alone or as part of a test battery to investigate the effects of lesion, genetic, or pharmacological manipulations. The task has also evolved to assess spatial memory in rodents via a modified version referred to as object placement or object location (referred to herein as object placement) [2]. As such, OR is used to assess memory for the identity of objects (i.e., “what”) and object placement (OP) is used to assess memory for the location of objects (i.e., “where”). To this end, OR in rodents is generally considered a non-spatial memory task involving the hippocampus, perirhinal, entorhinal, and parahippocampal cortices [3], [4], [5], whereas OP in rodents is considered a spatial memory task that relies primarily on the hippocampus [6]. However, the brain regions involved in OR in rodents and other species have been the subject of intense debate, particularly the role of the hippocampus in mediating OR. Although this issue is not the primary focus of this review, rodent data from our laboratory and others do support a role for the hippocampus in OR, as will be discussed below. Therefore, this review is written from the perspective that the hippocampus is essential for memory formation in both OR and OP. Because the amount of data collected on hormonal regulation of object memory in rodents far outnumbers the amount of data collected in other species, this review will limit discussion to studies employing rats or mice as subjects.
OR and OP are particularly well suited for investigating the molecular processes underlying the formation of hippocampal-dependent memories in rodents. First, they take advantage of a rodent's natural tendency to explore novel stimuli, while avoiding other potentially confounding variables. For example, no rule learning is required, nor are any rewarding or punishing stimuli involved that may influence motivational, rather than mnemonic, aspects of task performance [1], [7]. Therefore, memory can be measured in the absence of confounds due to the stress of nutrient restriction (as commonly used in the radial arm maze and T-maze), shock (as used in fear conditioning), or submersion in water (as used in the Morris water maze). Second, OR and OP are true one-trial learning tasks. This quality makes them ideal for studying the effects of acute drug or hormone treatments, which may be given pre- or post-training to investigate effects on different phases of learning and memory such as encoding, consolidation, and retrieval.
This unique combination of one-trial learning in a relatively stress-free environment has appealed in recent years to behavioral neuroendocrinologists seeking to identify the molecular mechanisms through which sex steroid hormones, such as 17β-estradiol (E2) and progesterone (P4), influence memory across the rodent lifespan. The low stress associated with OR and OP testing is advantageous for behavioral endocrinologists because corticosteroids released in response to more stressful tasks may interact with ovarian hormones and could confound the interpretation of results [8], [9]. Tasks like OR and OP that do not provoke a strong stress response allow for effects of ovarian hormones on memory to be more clearly identified in rodents. Furthermore, behavioral endocrinologists have found that E2 and P4 can very rapidly impact hippocampal function [10], [11], [12], [13], and therefore, the one-trial nature of OR and OP makes these tasks particularly useful for identifying the molecular mechanisms underlying hormonal regulation of memory consolidation. As such, both OR and OP have been widely used in the past decade to study the effects of sex steroid hormones on hippocampal learning and memory in rats and mice.
However, as will be seen below, investigators have taken very different approaches to studying hormonal regulation of object memory in rodents. Both rats and mice of various strains have been used, with studies employing mice generally outnumbering those using rats. Although species differences could influence the effects of hormones on OR, OR does not appear to differ between male rats and mice [14], and effects of E2 and P4 on OR and OP are remarkably consistent among rat and mouse studies. However, an interesting new report showing that the effects of estrogen receptor agonists on OR differ in two mouse strains suggests that strain or species may influence the receptor mechanisms through which hormones regulate object memory [15]. It is also important to note that although several studies have used gonadally intact females, the vast majority of studies to date administered exogenous hormones to ovariectomized females because removal of the ovaries eliminates the ovarian hormone fluctuations generated by the estrous cycle. In most studies, treatment was started at the time of ovariectomy or within a week afterward, but recent data show that long periods of ovarian hormone deprivation after ovariectomy eliminate the memory-enhancing effects of E2 on OR [16]. As such, timing of treatment relative to ovariectomy is an important variable for investigators to consider. Among exogenous hormone studies, most administered only E2. Therefore, much less is known about the effects of P4 on memory, alone or in combination with E2. Finally, a key difference among many studies is the timing of treatment relative to testing, as an increasing number of studies have administered E2 and P4 immediately after training to examine rapid effects of these hormones on object memory consolidation. In general, E2 or P4 given prior to, or immediately after, training enhance OR and OP, so the neural mechanisms underlying these effects are likely to be similar. As with any drug treatment, however, the effects of these hormones on object memory depend on many factors such as dose, route of administration, and age of subjects. Nevertheless, E2 and P4 generally enhance memory in the OR and OP tasks, as will be illustrated in the sections below.
The primary goal of this review is to survey literature examining the regulation of OR and OP in rats and mice by ovarian sex steroid hormones. To set the stage for this discussion, we will first address the role of the hippocampus in OR, provide an overview of ovarian hormone regulation of hippocampal function, and detail the protocols most commonly used in hormone studies. We will then describe the effects of natural estrous cycling and exogenous hormone administration on OR and OP. Finally, we will end by reviewing the molecular mechanisms known to mediate the memory-enhancing effects of E2 and P4 on OR and OP, and discuss how the reduction of ovarian hormone levels during aging affects memory in these tasks. Our intent is to highlight the importance of ovarian sex steroid hormones in regulating object memories in rodents and encourage investigators to consider the role that these hormones may play in their own experimental designs.
Section snippets
Role of the hippocampus in object recognition
Since their introduction in the 1980s and 1990s, one-trial OR and OP tasks have gained traction as standard tests for rodent memory in behavioral neuroscience and behavioral neuroendocrinology alike. Sensitive to hormones, aging, and drug treatments, OR and OP are most commonly associated with measures of episodic hippocampal memory. In addition to its role in other well known tasks such as the Morris water maze [17], contextual fear conditioning [18], and radial arm maze [18], the hippocampus
Effects of ovarian hormones in the hippocampus: A general overview
Although the contributions of the hippocampus to OR may be the subject of continued debate, an increasing body of research supports a role for the hippocampus in mediating OR memory consolidation. As a result, OR has become a useful tool for studying the effects of various neuromodulators, including sex steroid hormones, on hippocampal memory. To provide context for a discussion of this work, this section will describe the ways in which ovarian sex steroid hormones regulate hippocampal
OR and OP protocols typically used in behavioral endocrinology
Since their initial introduction for use in rats, OR and OP have been extended for use in mice and modified in variety of ways to suit the needs of different investigators and their experimental questions [1], [134], [135], [136]. Modifications include varying the number and size of the objects, size and shape of the testing arena, light levels in the testing arena, duration of habituation, and delay between training and testing. Although the experimental protocols for OR and OP differ somewhat
The estrous cycle
Studying the influence of endogenous E2 and P4 levels on hippocampal memory in gonadally intact females is inherently challenging in light of the naturally fluctuating hormones levels across the 4–5 day estrous cycle and the multi-day protocols typically used to test learning and memory. As such, very few studies have examined the effects of E2 and P4 on hippocampal memory in gonadally intact females. However, some investigators have capitalized on the one-trial nature of OR and OP to test the
Effects of ovariectomy on OR and OP
Given the effects of circulating ovarian hormones on OR and OP in gonadally intact rodents, one might ask whether ovariectomy itself has detrimental effects on memory in these tasks. Few studies have addressed this issue directly, but the data indicate that ovariectomy impairs memory in both tasks. One study of ovariectomized rats and gonadally intact sham-operated rats tested memory in the OR and OP tasks weekly for seven weeks after surgery, which was conducted at approximately 2.5 months of
Molecular mechanisms in the dorsal hippocampus underlying the beneficial effects of E2 and P4 on object memory
Numerous molecular mechanisms within the hippocampus are likely involved in the hormonal regulation of object memories. As discussed in Section 3 above, E2 and P4 may act via classical or non-classical mechanisms to alter hippocampal spinogenesis, neurogenesis, excitability, synaptic plasticity, cell signaling, epigenetic processes, and gene expression. Many of the rapid effects of these hormones on object memory have recently been attributed to non-classical actions of E2 and P4 (see below).
Regulation of object memory in aging females
In women, menopause occurs in the early 50s and is characterized by the cessation of reproductive function due to a sharp decline in circulating estrogens and progestins. In rodents, regular estrous cycling also declines in middle age due to age-related reductions in E2 and P4 levels, and the transition from normal to abnormal cycling has been associated with the onset of hippocampal memory decline [201], [202]. Ovarian hormones play a significant role in hippocampal neuroprotection [203], [204]
Conclusions and future directions
E2 and P4 are important regulators of object recognition and spatial memory in rodents, as illustrated by their effects on memory in the OR and OP tasks. Our laboratory and others have made great strides in recent years using these tasks as tools to understand the extent to which E2 and P4 regulate hippocampal memory formation. We have begun to identify the essential receptors, cell-signaling pathways, and epigenetic processes necessary for E2 and P4 to enhance OR and OP, but we have
Acknowledgements
The University of Wisconsin-Milwaukee supported this writing of this review. The empirical work from our laboratory described in this review was supported by grants from the National Institute on Aging (AG022525), National Institute of Mental Health (MH065460), Alzheimer's Association (IIRG-03–6051), American Federation for Aging Research/Pfizer (Grant in Hormones and Aging), and University of Wisconsin-Milwaukee (Research Growth Initiative Award) to K.M.F., an Ellison Medical Foundation/AFAR
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