Direct prion neuroinvasion following inhalation into the nasal cavity

Abstract

Inhalation of prions into the nasal cavity is an efficient route of infection. Following inhalation of infectious prions, animals develop disease with a similar incubation period compared with per os exposure, but with greater efficiency. To identify the reason for this increased efficiency, we identified neural structures that uniquely innervate the nasal cavity and neural structures known to mediate neuroinvasion following oral infection and used immunohistochemistry to determine the temporal and spatial accumulation of prions from hamster tissue sections containing cell bodies and axons at 2-week intervals following prion exposure. Prions were identified in the trigeminal ganglion, the spinal trigeminal tract in the brainstem, the intermediolateral cell column of the thoracic spinal cord, and the dorsal motor nucleus of the vagus/solitary nucleus complex months prior to detection of prions in the olfactory bulb or superior cervical ganglion. These results indicate that the trigeminal nerve, but not the olfactory nerve or sympathetic nerves, are involved in neuroinvasion following inhalation of prions into the nasal cavity. The detection of prions in the intermediolateral cell column of the thoracic spinal cord and dorsal motor nucleus of the vagus nerve 14 weeks following inhalation is consistent with inoculum crossing the alimentary wall and infecting the enteric nervous system via this route of infection. Neuroinvasion via the trigeminal nerve, in combination with entry into the central nervous system via autonomic innervation of the enteric nervous system, may contribute to increased efficiency of nasal cavity exposure to prions compared with per os exposure in hamsters.IMPORTANCEInhalation of prions into the nasal cavity is thought to be a route of infection in naturally acquired prion diseases. Experimental studies indicate that inhalation of prions is up to two orders of magnitude more efficient compared with ingestion. The mechanisms underlying this observation are poorly understood. We found a previously unreported direct route of neuroinvasion from the nasal cavity to the nervous system. Importantly, the peripheral ganglia involved may be a useful tissue to sample for prion diagnostics. Overall, identification of a new route of neuroinvasion following prion infection may provide an anatomical basis to explain the increased efficiency of infection following prion inhalation.

Keywords: nasal cavity; neuroinvasion; prion diagnostics; prion disease; trigeminal ganglia; trigeminal nerve.

Fig 1

Detection of PrP^Sc immunoreactivity (ir) following inhalation of prions into the nasal cavity (NC) at 22 weeks post-inoculation. There was robust PrP^Sc-ir in a subpopulation of neurons in the trigeminal (V) ganglion (panel A) following inhalation of infected brain homogenate (bh) but was not detected following inhalation of mock-infected bh (panel B). There was a small number of PrP^Sc-ir neurons in the superior cervical ganglion following inhalation of infected bh (indicated by arrow in panel C) and no detectable ir following inhalation of mock-infected bh (panel D). PrP^Sc-ir was pronounced in neurons in the intermediolateral cell column (ICC) of the thoracic spinal cord following inhalation (panel E); there was no detectable ir in the ICC of the spinal cord (panel F) when adjacent tissue sections were processed using an isotype control. The dorsal motor nucleus of the vagus and solitary nucleus contained PrP^Sc-ir following inhalation of infected bh; a medium power view shows widespread PrP^Sc-ir in the SN (panel G); PrP^Sc-ir was not detected when an isotope control antibody was utilized on an adjacent section. All scale bars = 50 µm.

Fig 2

PrP^Sc-ir in CN V ganglia and V structures in the brainstem increases over time following inhalation of prions. There was an increase in the number of PrP^Sc-ir CN V ganglion neurons and in the amount of intraneuronal PrP^Sc-ir from 10 weeks (panel A) to 14 weeks (panel C) to 18 weeks (panel E) following inhalation of infected bh. The same pattern of increased PrP^Sc-ir was seen in the spinal V tract and spinal V nucleus at 10 (panel B), 14 (panel D), and 18 weeks (panel F) following inhalation of infected bh. The insets in panels B, D, and F are enlarged views of the areas within the boxes showing punctate PrP^Sc depositions. The arrows in panels D and F indicate additional areas of PrP^Sc deposition following inhalation of infected bh. Scale bars in panels A, C, E = 50 µm; scale bars in panels B, D, F = 200 µm.

Fig 3

Detection of PrP^Sc in the ICC of the thoracic spinal cord and DMNV/SN following inhalation of infected bh into the NC. PrP^Sc-ir was initially restricted to the DMNV of the medulla (panel A) and to a small number of neurons in the ICC of the thoracic spinal cord (panel C) at 14 weeks following inhalation of infected bh. PrP^Sc-ir spread noticeably in the DMNV and SN and to adjacent areas (panel B) and spread throughout the gray matter of the thoracic spinal cord (panel D) at 18 weeks post-inhalation. Note that the brain/brainstem sections were cut in the sagittal plane; rostral is toward the top of the figures, and the dorsal surface of the medulla is to the right. The solitary nucleus was identifiable as a relatively narrow tubular structure elongated in the rostral-caudal plane, ventral to the nucleus gracilis (NG), and dorsal to the DMNV. Insets in panels A and C are enlarged views of the areas within the boxes showing the relatively modest amount of PrP^Sc-ir at 14 weeks post-inhalation. Scale bars = 200 µm.