Robert D. Hawkins

Control of Memory Formation Through Regulated Expression of a CaMKII Transgene

One of the major limitations in the use of genetically modified mice for studying cognitive functions is the lack of regional and temporal control of gene function. To overcome these limitations, a forebrain-specific promoter was combined with the tetracycline transactivator system to achieve both regional and temporal control of transgene expression. Expression of an activated calcium-independent form of calcium-calmodulin-dependent kinase II (CaMKII) resulted in a loss of hippocampal long-term potentiation in response to 10-hertz stimulation and a deficit in spatial memory, a form of explicit memory. Suppression of transgene expression reversed both the physiological and the memory deficit. When the transgene was expressed at high levels in the lateral amygdala and the striatum but not other forebrain structures, there was a deficit in fear conditioning, an implicit memory task, that also was reversible. Thus, the CaMKII signaling pathway is critical for both explicit and implicit memory storage, in a manner that is independent of its potential role in development.

Long-term potentiation is reduced in mice that are doubly mutant in endothelial and neuronal nitric oxide synthase.

Nitric oxide (NO) has been implicated in hippocampal long-term potentiation (LTP), but LTP is normal in mice with a targeted mutation in the neuronal form of NO synthase (nNOS-). LTP was also normal in mice with a targeted mutation in endothelial NOS (eNOS-), but LTP in stratum radiatum of CA1 was significantly reduced in doubly mutant mice (nNOS-/eNOS-). By contrast, LTP in stratum oriens was normal in the doubly mutant mice. These results provide the first genetic evidence that NOS is involved in LTP in stratum radiatum and suggest that the neuronal and endothelial forms can compensate for each other in mice with a single mutation. They further suggest that there is also a NOS-independent component of LTP in stratum radiatum and that LTP in stratum oriens is largely NOS independent.

Impairment of spatial but not contextual memory in CaMKII mutant mice with a selective loss of hippocampal ltp in the range of the θ frequency

We assessed hippocampal-dependent memory in mice with a Ca(2+)-independent form of CaMKII generated by the introduction of an aspartate at amino acid 286. The CaMKII-Asp-286 mice show normal LTP at high frequency stimulation, but in the 5-10 Hz range, they show a shift in the frequency-response curve favoring LTD. This range of frequencies is similar to the theta rhythm, which is associated with exploration in rodents. Using the Barnes maze to assess spatial memory, we found the transgenic mice could not learn to navigate to a specific location using spatial cues. In contrast, one line of transgenic mice performed normally in contextual fear conditioning, a task that is also hippocampal dependent. This dissociation between spatial and contextual memory suggests that even though both require the hippocampus, they may be mediated by different synaptic mechanisms.

IS HETEROSYNAPTIC MODULATION ESSENTIAL FOR STABILIZING HEBBIAN PLASTICITY AND MEMORY

In 1894, Ramón y Cajal first proposed that memory is stored as an anatomical change in the strength of neuronal connections. For the following 60 years, little evidence was recruited in support of this idea. This situation changed in the middle of the twentieth century with the development of cellular techniques for the study of synaptic connections and the emergence of new formulations of synaptic plasticity that redefined Ramón y Cajals idea, making it more suitable for testing. These formulations defined two categories of plasticity, referred to as homosynaptic or Hebbian activity-dependent, and heterosynaptic or modulatory input-dependent. Here we suggest that Hebbian mechanisms are used primarily for learning and for short-term memory but often cannot, by themselves, recruit the events required to maintain a long-term memory. In contrast, heterosynaptic plasticity commonly recruits long-term memory mechanisms that lead to transcription and to synaptic growth. When jointly recruited, homosynaptic mechanisms assure that learning is effectively established and heterosynaptic mechanisms ensure that memory is maintained.

Inducible and reversible gene expression with the rtTA system for the study of memory.

To obtain rapidly inducible and reversible expression of transgenes in the forebrain of the mouse, we have combined the reverse tetracycline-controlled transactivator (rtTA) system with the CaMKIIalpha promoter. We show that doxycycline induces maximal gene expression in neurons of the forebrain within 6 days and that this expression can be reversed by removal of doxycycline. Using calcineurin as a test transgene, we show that doxycycline-induced expression impairs both an intermediate form of LTP (I-LTP) in the hippocampus and the storage of spatial memory. The reversibility of the rtTA system in turn allowed us to examine the effects of the transgene on memory retrieval after normal storage had occurred. This examination suggests that retrieval requires some of the same molecular components required for storage.

Mice Expressing Activated CaMKII Lack Low Frequency LTP and Do Not Form Stable Place Cells in the CA1 Region of the Hippocampus

To relate different forms of synaptic plasticity to the formation and maintenance of place cells in the hippocampus, we have recorded place cells in freely behaving, transgenic mice that express a mutated Ca2+-independent form of CaM Kinase II. These mice have normal long-term potentiation (LTP) at 100 Hz, but they lack LTP in response to stimulation at 5-10 Hz and are impaired on spatial memory tasks. In these transgenic mice, the place cells in the CA1 region have three important differences from those of wild types: they are less common, less precise, and less stable. These findings suggest that LTP in the 5-10 Hz range may be important for the maintenance of place-field stability and that this stability may be essential for the storage of spatial memory.

A Comparative Analysis of the Molecular Mechanisms Contributing to Implicit and Explicit Memory Storage in Aplysia and in the Hippocampus

Studies of implicit and explicit memory suggest that modulation of synaptic strength and structure is a fundamental mechanism by which memories are encoded, processed, and stored within the brain. Two model systems have been extensively studied as examples of these two forms of memory: sensitization in the marine snail Aplysia californica as an example of implicit memoryxa0and spatial memory formation in rodents as an example of explicit memory. In this chapter, we discuss and compare critical synaptic sites and cellular and molecular mechanisms of implicit and explicit memory storage, and describe new approaches to relating these mechanisms to behavior.

Presynaptic Mechanisms of Plasticity and Memory in Aplysia and Other Learning-Related Experimental Systems

Cellular studies of memory suggest that experience-dependent synaptic plasticity is a fundamental mechanism by which memories are encoded, processed, and stored within the brain. In this chapter, we focus on recent advances in our understanding of the molecular mechanisms of a simple form of learning-related plasticity, presynaptic facilitation, that contributes to short-term, intermediate-term, and long-term forms of memory in the marine invertebrate Aplysia californica . We also briefly review mechanisms of presynaptic plasticity in other experimental systems, which share many similarities with those in Aplysia and may play important roles in memory as well.