Regulation of sympathetic tone and arterial pressure by rostral ventrolateral medulla after depletion of C1 cells in rat

The rostral ventrolateral medulla (RVLM) is critical for maintaining basal sympathetic vasomotor tone and is an essential component of many sympathetic reflexes (Reis et al. 1989; Stornetta et al. 1989; Guyenet, 1990; Verberne & Guyenet, 1992; Koshiya et al. 1993; Dampney, 1994a; Sun, 1995; McCulloch et al. 1999). The C1 adrenergic neurones coincide with a region of the RVLM that regulates sympathetic nerve activity (SNA) and arterial pressure (AP) (Ross et al. 1981, 1984b; Reis et al. 1989), and these cells have been proposed to be important for the generation of sympathetic vasomotor tone and AP. The spinal axons of many C1 cells target sympathetic preganglionic neurones or their immediate vicinity (Ross et al. 1984a; Hokfelt et al. 1984; Milner et al. 1988; Jansen et al. 1995a, b). In addition, many bulbospinal C1 neurones are inhibited by increased AP (Lipski et al. 1995; Schreihofer & Guyenet, 1997; Verberne et al. 1999), a hallmark of the sympathetic vasomotor efferents that regulate blood pressure and cardiac output. However, the C1 cells are not the only efferent projection of the RVLM towards the thoracic spinal cord. This structure also contains other highly active, bulbospinal neurones that are inhibited by stimulation of arterial baroreceptors, but do not contain tyrosine hydroxylase (TH) or phenylethanolamine-N-methyltransferase (PNMT) (Lipski et al. 1995; Schreihofer & Guyenet, 1997). The properties of these lightly myelinated, non-catecholaminergic cells (Lipski et al. 1996) suggest that they could be a source of supraspinal glutamatergic drive to sympathetic preganglionic neurones (Deuchars et al. 1995), which is essential for the generation of resting sympathetic tone and AP (Huangfu et al. 1994). The relative contributions of the C1 cells and the non-catecholaminergic RVLM neurones to the generation of sympathetic vasomotor tone are not known. One way to address this question would be to identify deficits in the regulation of sympathetic tone that result from the selective destruction of one or the other cell type. Until recently, neither cell type could be targeted because C1 neurones are insensitive to the classic catecholaminergic neurotoxin 6-hydroxydopamine (6-OHDA; Jonsson et al. 1976), and a specific marker to target the non-catecholaminergic neurones has not been identified. However, a newly introduced immunotoxin produced by conjugating the ribosomal toxin saporin to an anti-dopamine-β-hydroxylase antibody (anti-DβH-SAP) now provides the means to lesion C1 cells while sparing non-catecholaminergic neurones in the RVLM (Wrenn et al. 1996). Membrane-bound DβH is exteriorized during exocytosis and acts as a specific receptor for the internalization of anti-DβH-SAP into noradrenergic and adrenergic neurones. Once inside, saporin blocks protein synthesis to cause the death and eventual elimination of the cell (Stripe & Barbieri, 1986). Anti-DβH-SAP has proved capable of selectively lesioning C1 neurones within the RVLM after administration into terminal fields or next to cell bodies (Wrenn et al. 1996; Madden et al. 1999; Schreihofer & Guyenet, 2000). We have previously shown that the depletion of bulbospinal C1 cells by intraspinal microinjection of anti-DβH-SAP does not chronically alter AP or abolish sympathetic vasomotor tone in anaesthetized rats (Schreihofer & Guyenet, 2000). However, the sympathoexcitatory response to cyanide is virtually eliminated (Schreihofer & Guyenet, 2000). In the present study we examined whether the RVLM continues to maintain basal sympathetic tone after depletion of bulbospinal C1 cells with intraspinal microinjection of anti-DβH-SAP. Because this treatment also depletes bulbospinal pontine noradrenergic neurones (Schreihofer & Guyenet, 2000), we also examined the effects of selective depletion of these cells by pontine microinjection of 6-OHDA. Furthermore, we characterized the remaining barosensitive bulbospinal RVLM neurones to determine whether the lightly myelinated non-catecholaminergic cells could be maintaining SNA. Finally, we determined whether the reduced sympathoexcitatory responses in rats treated with anti-DβH-SAP could be the result of a deficit in the ability of the RVLM to activate SNA.