Location-specific activation of the paraventricular nucleus of the hypothalamus by localized inflammation

Abstract

The existence of an immunological homunculus has been proposed, but evidence for location-specific response of the central nervous system to immunological stimulation is lacking. In this study, we show that inflammation induced by injection of casein into one of the causes c-fos expression in the paraventricular nucleus of the hypothalamus (PVN) in an asymmetrical manner: much stronger activation is always induced in the contralateral PVN. Unilateral sciatic nerve transection abolished the casein-induced PVN activation if casein was injected into the hindlimb with the nerve transection, but had no effect if casein was injected into the hindlimb with intact nerve innervation. Injection of casein into one the forelimbs also caused contralateral PNV activation. Further, stronger PVN activation was found in the anterior PVN after the forelimb injection, but in the posterior PVN after the hindlimb injection. Casein-induced PVN activation is absent in IL-1R1 KO, IL-6 KO, TNFα KO, and in C3H/HeJ (TLR4 mutant) animals. In comparison, injection of LPS, a systemic inflammagen, into one hindlimb induced bilateral PVN activation but injection of live Escherichia coli into one hindlimb induced contralateral PVN activation. These results support the notion that local inflammation may activate the PVN by neural routes in a location-specific manner.

Figure 1

Representative low-magnification microphotographs show labeling of c-fos (A-C) in coronal sections containing PVN. A: home cage control. B: c-fos expression 6 h after the saline injection into the right hindlimb muscle. C: c-fos expression 6 h after casein injection into the right hindlimb. Arrows point to c-fos positive cells induced by both casein and saline injections; arrowheads point to c-fos positive cells induced by the casein injection. * indicates the pinhole that marks the left side of the brain.

Figure 2

Representative high-magnification microphotographs show labeling of c-fos in coronal sections containing PVN after intramuscular casein injection into the right hindlimb (A–C) or into the left hindlimb (a–c). Animals were sacrificed at 4 (A,a), 6 (B,b), or 10 (C,c) h after the casein injections.

Figure 3

Number of c-fos positive cells in the ipsilateral (iPVN) vs. contralateral PVN (cPVN, relative to the injection site). * indicates statistical significance (p<0.05, iPVN vs. cPVN).

Figure 4

Representative low-magnification microphotographs show labeling of c-fos (A–C) in coronal sections containing PVN. All animals received left sciatic nerve transection and were sacrificed 6 h after intramuscular injections. A: saline was injected into the right hindlimb. B: saline was injected into the left hindlimb. C: casein was injected into the right hindlimb. D: casein was injected into the left hindlimb. Arrow points to c-fos positive cells in the PVN; arrowheads point to c-fos positive cells in the SON.

Figure 5

Representative high-magnification microphotographs show labeling of c-fos in coronal sections containing PVN after intramuscular casein injection into the right forelimb (A–F) or into the right hindlimb (a-f). Serial sections in the anterior to posterior order from two animals are shown. aPVN: anterior PVN; pPVN: posterior PVN.

Figure 6

Quantitative analysis of the number of c-fos positive cells in the left PVN after casein injection into the right forelimb or the right hindlimb. * indicates statistical significance (p<0.05, aPVN vs. pPVN).

Figure 7

Representative high-magnification microphotographs show labeling of c-fos in transverse sections of the spinal cord after intramuscular casein injection into the left forelimb (A–B) or into the left hindlimb (C–D). Dye on the edge marks the side of casein injection. Arrows point to c-fos positive cells in the dorsal horn in the ipsilateral spinal cord (A and C). No c-fos induction was found in the contralateral spinal cord (B and D). A and B were taken from spinal level C5-TH1; C and D were taken from spinal level L2-S2.

Figure 8

Representative microphotographs show c-fos labeled sections containing various supraspinal brain regions in both left (L) and right (R) sides of the brain. Animals were sacrificed 6 h after casein was injected into the right hindlimb muscles. The scale bar in C shows magnification level for pictures A-I. The magnification level in J is marked by its own scale bar. VLM: ventrolateral medulla; PB: parabrachial nucleus; AP: area postrema; NTS: nucleus of the solitary tract; LC: locus coeruleus; CeA: central nucleus of the amygdala; BST: bed nucleus of the stria terminalis; AH: anterior hypothalamus.

Figure 9

Representative high-magnification microphotographs show c-fos labeled coronal sections containing PVN after intramuscular casein injection into the right hindlimb of C3H/HeJ (A), IL-1R1 KO (B), IL-6 KO (C), and TNFα KO (D) animals.

Figure 10

Representative microphotographs show c-fos labeled coronal sections containing PVN (A and B), SFO (C and D), and AP/NTS (E and F) after intramuscular injection into the right hindlimb of LPS (left panels), or live E. coli (right panels). COX-2 labeled sections are also shown (H and I).