Divide and conquer: an L-type voltage-gated calcium channel subtype finds a role in conditioned fear.

The function of L-type voltage-gated calcium channels (LVGCCs) in learning and memory has been somewhat elusive. For some time we have known that there is progressive decrease in learning and memory with age, and a number of researchers have argued that this is because of a gradual increase in LVGCCs through the life cycle (Barad 2003). Consistent with this observation, LVGCC inhibitors of various types have been observed to improve learning in aged, but not in young animals. Of course, these observations are inconsistent with a role for LVGCCs in promoting learning. Similarly, the search for an LVGCC-dependent type of longterm potentiation (LTP) has been somewhat frustrating. Since calcium entry into cells is an essential step in many forms of synaptic plasticity including LTP, and since such synaptic plasticity is presumed to be a mechanism underlying learning, it seems reasonable to presume that LVGCCs provide one means of calcium entry contributing to learning. However, a number of labs have found that LVGCC-dependent LTP only occurs in the hippocampus when NMDA receptors are blocked, and when the rate of pathway stimulation is extremely high—at least 200 Hz (Barad 2003). This is an unlikely scenario in vivo. However, in the amygdala a more physiological pairing of postsynaptic depolarization with presynaptic stimulation of the thalamo–amygdala pathway has been shown to induce an LVGCC-dependent LTP (Weisskopf et al. 1999; Bauer et al. 2002). Other protocols also generate LVGCC-dependent LTP in various amygdala pathways (Lee et al. 2002; Niikura et al. 2004; Fu and Shinnick-Gallagher 2005; Drephal et al. 2006). Thus, the amygdala appears to be a particularly promising location to support for LVGCC-dependent learning. Regardless of whether physiologically plausible stimulation patterns can generate LVGCC-dependent LTP, the real test of whether these channels play an important role in learning can only come from behavioral tests. In fact, behavioral tests in the past few years have begun to demonstrate a potential role for LVGCCs in several forms of learning, especially using classically conditioned fear, which is well known to depend on the amygdala. Classical, or Pavlovian, conditioned fear of a neutral stimulus, like a tone or the context of conditioning (conditioned stimulus, CS), is created by pairing the CS with an intrinsically aversive unconditioned stimulus (US), typically a footshock. A single pairing of this kind generates robust conditioned fear of the CS that can last a lifetime (Gale et al. 2004). Behavioral scientists measure this fear in increased heart rate, blood pressure, and other physiological changes, as well as in a number of behavioral responses including fear-potentiated startle and freezing, the absence of all movement except breathing—the measure used in the paper from McKinney and Murphy (2006). Conditioned fear can be suppressed or decreased by the repeated presentation of the CS without any US, a process called extinction. Fear extinction learning shows complete LVGCC dependence, as no extinction occurs when training is performed in the presence of LVGCC inhibitors (Cain et al. 2002; Suzuki et al. 2004). An LVGCC inhibitor infused into the basolateral amygdaloid complex interferes with the consolidation phase of fear conditioning itself (Bauer et al. 2002). That is, freezing to the CS is equal to that of controls immediately after fear conditioning in the presence of LVGCC inhibition, but fear is substantially decreased compared with controls when tested 24 h later. These distinct roles of LVGCCs in fear conditioning and extinction beg for more thorough dissection, and McKinney and Murphy (2006) provide it. McKinney and Murphy (2006) provide an elegant example of how genetically modified mice can be used to clarify a confusing field and jump its dissection to a new level of detail. There are two major subtypes of LVGCC in the forebrain, CaV1.3 and CaV1.2. Since available pharmacological inhibitors do not distinguish between LVGCC subtypes, the dissection of the role of LVGCC subtypes in the brain can only be achieved by genetic means. Such a means recently became available though the generation of mice with genetic inactivation of the pore-forming subunit of one or the other LVGCC subtype, knocking out the function of the channel (Moosmang et al. 2005b). Studies using the CaV1.2 knockout mice demonstrated that this mutation disrupted NMDA-independent late LTP in the mossy fiber–CA1 pathway of the hippocampus (using a 200-Hz tetanus) and disrupted spatial learning (Moosmang et al. 2005a). McKinney and Murphy (2006) take advantage of the other LVGCC knockout, of CaV1.3, to examine the role of this subtype in fear learning. Although the CaV1.3 knockout mice are deaf, the authors had the luck to find that they are otherwise grossly intact neurologically and behaviorally. Their performance is equivalent to wild-type littermate controls in the rotarod test of vestibular function and reaction speed. It is also equivalent in open-field measures of activity and general anxiety. Thus satisfied with the general neurological and behavioral functioning of the CaV1.3 knockout mice, the authors proceeded to their main interest in the function of LVGCCs in fear learning. Again, because the CaV1.3 knockout mice are deaf, and because most cue fear conditioning depends on auditory stimuli, the authors elected to focus on contextual rather than cue fear. To generate contextual conditioned fear, the authors placed mice in a novel context for an acclimation period and then administered a single mild footshock. This treatment induces fear of the context measured by freezing when the animals are returned there. Comparing CaV1.3 knockout mice to wild-type littermates, the authors found that the CaV1.3 LVGCC subtype is crucial for the consolidation, but not the initial formation of conditioned fear of context, as CaV1.3 knockout mice showed fear equal to their littermates at 1 and 6 h after fear conditioning, but showed significantly less fear than the littermates at 24 h after conditioning. In a satisfying convergence, the time course McKinney and Murphy (2006) found for the dependence of conditioned fear on E-mail mbarad@mednet.ucla.edu; fax (310) 206-5855. Article is online at http://www.learnmem.org/cgi/doi/10.1101/lm.402806. Commentary