The olfactory nerve has a role in the body temperature and brain cytokine responses to influenza virus

Abstract

Mouse-adapted human influenza virus is detectable in the olfactory bulbs of mice within hours after intranasal challenge and is associated with enhanced local cytokine mRNA and protein levels. To determine whether signals from the olfactory nerve influence the unfolding of the acute phase response (APR), we surgically transected the olfactory nerve in mice prior to influenza infection. We then compared the responses of olfactory-nerve-transected (ONT) mice to those recorded in sham-operated control mice using measurements of body temperature, food intake, body weight, locomotor activity and immunohistochemistry for cytokines and the viral antigen, H1N1. ONT did not change baseline body temperature (Tb); however, the onset of virus-induced hypothermia was delayed for about 13 h in the ONT mice. Locomotor activity, food intake and body weights of the two groups were similar. At 15 h post-challenge fewer viral antigen-immunoreactive (IR) cells were observed in the olfactory bulb (OB) of ONT mice compared to sham controls. The number of tumor necrosis factor alpha (TNFalpha)- and interleukin 1beta (IL1beta)-IR cells in ONT mice was also reduced in the OB and other interconnected regions in the brain compared to sham controls. These results suggest that the olfactory nerve pathway is important for the initial pathogenesis of the influenza-induced APR.

Fig. 1

Photomicrographs of sagittal sections of the distal olfactory nerve and its connectivity with the olfactory bulb. (A) A sagittal section showing the axons of the olfactory receptor neurons connecting with the glomerular layer of the olfactory bulb in a mouse that received a sham surgery. (B) A sagittal section showing the loss of connectivity between the olfactory nerve axons and the olfactory bulb after the olfactory nerve transection surgery. OE, olfactory epithelium; ON, olfactory nerve; GL, glomerular layer. The scale bar is 0.02 mm.

Fig. 2

Time course of body temperature changes in olfactory nerve transected [ONT (closed circles)] or sham (open circles) mice following intranasal inoculation with influenza virus. The mice with a transected olfactory nerve (ONT) delay their hypothermia response to influenza virus by 13 h. Data points represent the means ± SEM at 1 h intervals. Asteriks (*) designate significant differences, p<0.05 between the ONT and the sham groups. The solid horizontal bars along the abscissa depict the 12 h dark periods.

Fig. 3

Locomotor activity responses to PR8 challenge in ONT (closed circles) or sham (open circles) mice. Data points represent the means ± SEM at 1 h intervals. No significant differences were observed between the olfactory nerve transected (ONT) and the sham groups. After live viral challenge, locomotor activity increased for about 4 h compared with the same time of the baseline day. It then decreased below baseline values in both groups. The second day after viral challenge, activity remained below baseline levels. The solid horizontal bars along the abscissa mark the 12 h dark periods.

Fig. 4

Food intake and body weight following intranasal inoculation with influenza virus in ONT or sham mice. Food intake (A) and body weight (B) were determined for 6 days. (A) Data points represent the food consumption for the 24 h prior to the measurement. A decrease in food intake was observed in the ONT (closed circles) and sham (open circles) groups that received live virus compared to the sham mice that received the boiled virus (open triangles). (B) Data points represent the percentage of body weight in relationship to the initial body weight (day of virus challenge or day 0). Mice inoculated with live virus (ONT and sham) lost weight when compared with the sham mice that received boiled virus. Asterisks (*) represent significant differences, p<0.05 between the ONT and the sham groups compared to the boiled virus group.

Fig. 5

Photomicrographs of olfactory bulb coronal sections from infected ONT or sham mice killed 15 h PI and stained for influenza H1N1. (A) Coronal section from an ONT mouse that received live PR8, with little evident H1N1 immunoreactivity is evident. (B) A higher magnification of the GL shown in (A). (C) Coronal section from a sham mouse showing the H1N1 immunoreactivity in the ON and the GL. (D) A higher magnification of the GL in (C). Arrows point to some stained cells with glial-like morphology. The scale bar for lower magnification pictures (A and C) is 0.02 mm; for higher magnification pictures (B and D) the scale bar is 0.025 mm. ML, mitral cell layer; EPL, external plexiform layer; GL, glomerular layer; ON, olfactory nerve.

Fig. 6

Quantitative analyses of the number of the viral antigen H1N1-immunoreactive (IR) cells in the OB of ONT (n=6) or sham (n=6) mice inoculated with live virus. The number of H1N1-IR cells was significantly different between the two groups. Mice were inoculated with virus 10 days subsequent to olfactory nerve transection. Asterisk (*) indicates p<0.05