Cerebellar norepinephrine modulates learning of delay classical eyeblink conditioning: evidence for post-synaptic signaling via PKA.

The neurotransmitter norepinephrine (NE) is strongly implicated in cerebellar-dependent learning and memory in adaptation to a patterned rod runway task (Watson and McElligott 1983; Bickford 1995), in adaptation of the vestibulo-ocular reflex gain (VOR) (Pompeiano et al. 1991), and in cerebellar-dependent delay classical eyelid conditioning (Gould 1998; McCormick and Thompson 1982; Winsky and Harvey 1992; Cartford et al. 2002). The delay form of classical eyelid conditioning is a valuable model for studying NE because the functional anatomy is so well characterized and localization of learning in the cerebellum is strongly supported by the literature. Neurons within cerebellar lobule HVI and the interpositus nucleus (IP) in rabbits and rats show conditioning-related activity (Berthier and Moore 1986; Gould and Steinmetz 1994; Rogers et al. 2001). Classical conditioning of the eyelid response is disrupted by lesions of cerebellar lobule HVI (Yeo et al. 1985; Steinmetz and Sengelaub 1992; Nordholm et al. 1993; Perrett et al. 1993) and is abolished by lesions of cerebellar IP (Clark et al. 1992, 1997; Krupa et al. 1993; Clark and Lavond 1996; Rogers et al. 2001). However, Purkinje cell degeneration mutant (pcd) mice acquire the eyelid response (Chen et al. 1996), and both pcd mice as well as the jaundiced Gunn rat (also a mutant with loss of Purkinje cells) have normal to elevated levels of NE innervation and functional activity in cerebellar cortex and deep nuclei after degeneration of the Purkinje cell layer (Ghetti 1981; Kostrozewa and Harston 1986; Onozuka et al. 1990). Norepinephrine is known to modulate the action of other neurotransmitters in both the cerebellar cortex and the deep nuclei (Gould et al. 1997) and can amplify afferent inputs to cerebellar Purkinje neurons. This effect is mediated through the β-noradrenergic receptor (Yeh and Woodward 1983; Woodward et al. 1991). Noradrenergic receptor activation signals a G-protein-coupled signal transduction cascade in which adenyl cyclase (AC), cyclic-adenosine-monophosphate (cAMP), and protein kinase A (PKA) are activated and lead to the downstream phosphorylation of multiple substrates including cAMP-responsive element binding protein (CREB). Outside of the cerebellum, cAMP, PKA, and phosphorylated CREB (pCREB) have been implicated in the establishment of synaptic changes necessary for both short-term and long-term memory formation (Taylor et al. 1999; Muller 2000; Vianna et al. 2000; Baldwin et al. 2002; Shobe 2002), and studies in long-term potentiation (LTP) and long-term depression (LTD) support these behavioral findings (Huang et al. 1994; Huang and Kandel 1996; Nayak et al. 1998; Rotenberg et al. 2000). In eyelid conditioning, in particular, Chen and Steinmetz (2000) have shown that localized blocking of a range of kinase activity disrupts acquisition but not retention of conditioning in rabbits. Genetic expression in cerebellum related to eyelid conditioning has not been studied, but there is accumulating evidence for gene expression relative to learning in hippocampus (Donahue et al. 2002) and that blocking genetic expression in cerebellum prevents acquisition of conditioned responses (Gomi et al. 1999). Our hypothesis is that the activity of NE at the β-noradrenergic receptors in cerebellum facilitates learning. Finally, we hypothesize that NE signaling mechanisms contribute to learning through activation of PKA, and therefore blocking the activation of PKA will result in learning deficits.