Given the extent of co-localization of NOS and VAChT in the LDT, we postulate that many of the NOS cells expressing the 5HT1A receptor in this structure are also cholinergic (Grant and Highfield, 1991; Kayama and Ogawa, 1987; Leonard et al

Given the extent of co-localization of NOS and VAChT in the LDT, we postulate that many of the NOS cells expressing the 5HT1A receptor in this structure are also cholinergic (Grant and Highfield, 1991; Kayama and Ogawa, 1987; Leonard et al., 1995; Satoh el al., 1983). 90%). The findings suggest this assembly of cells is a rostral extension of the LDT, one that it is subject to regulation by 5HT release. As such the present study suggests a link between 5HT signaling, activation of cholinergic/NOS neurons, and the stress response including Ly6a the pathophysiology underlying anxiety and depression. Bonferroni tests revealed that VAChT and VAChT-NOS expression in the caudal LW C LDT transition zone were significantly LDN193189 HCl different than all other DRN sub-regions (Figure 3; p .001). Open in a separate window Figure 1 Horizontal Sections of NOS and VAChT Staining across the DRNHorizontal sections of the rat brainstem at the level of the DRN and lateral dorsal tegmental nucleus (LDT) reveal the lateral wing sub-region of the DRN and the LDT (A, white box) form LDN193189 HCl a continuous column of cells along the rostral-caudal dimension. The caudal region of the LDT is bordered by the fourth ventricle (top of figure A and A). Tryptophan hydroxylase (TrpH) positive neurons (B, green) clearly intermingle with NOS positive cells (B, red) along the caudal lateral wing and rostral LDT border (B, B, B, white dashed line). Scale bars: A = 500 m, B = 250 m. Abbreviations in A: 3-oculomotor nucleus, 3V- third ventricle, 4- trochlear nucleus, 4n- trochlear nerve root, 4V- fourth ventricle, CnF- cuneiform nucleus, DpMe- deep mesencephalic nucleus, DR- dorsal raphe nucleus, dtg-dorsal tegmental bundle, EW- nucleus of Edinger Westphal, fr- fasciculus retroflexus, LDTg- lateral dorsal tegmental nucleus, mlf- medial longitudinal fasciculus, PF- parafascicular thalamic nucleus, RI- rostral interstitial nucleus of the mlf. Open in a separate window Figure 2 Juxtaposition of NOS, TrpH, and VAChT in the lateral wing of the DRNNOS-positive, tryptophan hydroxylase (TrpH)-negative cells located within the caudal portion of the DRN lateral wing (A, A) express the vesicular acetylcholine transporter (VAChT). High magnification images (B C B?, arrowheads) show NOS-positive neurons (B, red) co-localize with VAChT (B?, blue) but not TrpH (B, green). This finding indicates that NOS-positive, 5HT-negative cells within the DRN lateral wing are actually the rostral-most extension of the LDN193189 HCl cholinergic lateral dorsal tegmental nucleus. Scale bars: A = 250 m, A = 50 m, B = 30 m. Aq- cerebral aqueduct, MLF- medial longitudinal fasciculus. Open in a separate window Figure 3 Distribution and cell counts of NOS, TrpH, and LDN193189 HCl VAChT cells across the DRNThe average number of cells expressing tryptophan hydroxylase (TrpH, black bars), nitric oxide synthase (NOS, white bars), vesicular acetylcholine transferase (VAChT, diagonal bars), or NOS and VAChT (hashed bars) are compared caudal to rostral across the rostral LDN193189 HCl LDT/caudal LW border (?8.72 to ?8.30 Bremga) through to the rostral LW subregion of the DRN (?7.80 to ?7.30 Bregma). The LDT/LW border region (left side data bars; ?8.72 – ?8.30 Bregma) contains significantly higher numbers of VAChT and NOS-VAChT expressing cells than more rostral regions of the LW or any DRN midline subregions. (* p .001) Table 1 Cell counts per DRN sub-region at 40X magnification. Bonferroni tests indicated significant differences between several sub-regions for NOS, 5HT1A, and double labeled cells. Expression of 5HT1A receptor-positive cells was significantly higher in the intermediate ventromedial (iVM) sub-region compared to the intermediate LW (iLW; p .05), and, as expected, higher numbers of NOS cells were present in the caudal versus rostral LW sub-regions (p .05). The caudal LW C LDT transition zone had higher numbers of double labeled.