Measurement of Tactile Allodynia in a Murine Model of Bacterial Prostatitis

Summary

Infection of the prostate may be a contributing factor in mediating pelvic pain in chronic prostatitis. We describe the procedure for preparation of standardized bacterial inoculum, instillation of bacteria into the urethra of male mice and methodology for measuring tactile allodynia in mice over time.

Abstract

Uropathogenic Escherichia coli (UPEC) are pathogens that play an important role in urinary tract infections and bacterial prostatitis¹. We have recently shown that UPEC have an important role in the initiation of chronic pelvic pain², a feature of Chronic prostatitis/Chronic pelvic pain syndrome (CP/CPPS)^3,4. Infection of the prostate by clinically relevant UPEC can initiate and establish chronic pain through mechanisms that may involve tissue damage and the initiation of mechanisms of autoimmunity⁵.

A challenge to understanding the pathogenesis of UPEC in the prostate is the relative inaccessibility of the prostate gland to manipulation. We utilized a previously described intraurethral infection method⁶ to deliver a clinical strain of UPEC into male mice thereby establishing an ascending infection of the prostate. Here, we describe our protocols for standardizing the bacterial inoculum⁷ as well as the procedure for catheterizing anesthetized male mice for instillation of bacteria.

CP/CPPS is primarily characterized by the presence of tactile allodynia⁴. Behavior testing was based on the concept of cutaneous hyperalgesia resulting from referred visceral pain^8-10. An irritable focus in visceral tissues reduces cutaneous pain thresholds allowing for an exaggerated response to normally non-painful stimuli (allodynia). Application of normal force to the skin result in abnormal responses that tend to increase with the intensity of the underlying visceral pain. We describe methodology in NOD/ShiLtJ mice that utilize von Frey fibers to quantify tactile allodynia over time in response to a single infection with UPEC bacteria.

Protocol

1. Bacteria Preparation for Mouse Inoculation The following must be done under aseptic conditions. Take one 17 x 100 mm polypropylene tubes with caps and add 3 ml of fresh Luria broth (LB) media. Using autoclaved tips, take some frozen bacteria glycerol stock of strain CP12 and transfer a small quantity of the culture into the tube containing the LB media. Replace the tube cap so that oxygen is still able to enter the tube – the culture needs to grow under aerobic con…

Representative Results

We examined NOD/ShiLtj and C57BL/6J male mice for the presence of chronic pain upon bacterial infection. Male mice (5 to 7 weeks old) were instilled with saline or bacteria into the urethra (Figure 1). Mechanical stimulation of the pelvic area with von Frey filaments of saline-instilled C57BL/6J mice and UPEC-infected C57BL/6J mice resulted in a response frequency that did not change during the 28-day course of the experiment (Figure 1). In contrast, UPEC-infected NOD mice exhibited resp…

Discussion

Infection of the mouse prostate with UPEC allows for the in vivo modeling of events that may be involved in the pathogenesis of bacterial prostatitis, CP/CPPS or as a predisposing event in chronic inflammation. The methods described for bacterial preparation and instillation draw upon a large body of literature on UPEC models in female urinary tract infection^7,14. The model has wide applicability for studying pathogenesis, testing potential vaccine candidates and mechanisms of immune modulation. The a…

Materials

Name of the reagent	Company	Catalogue number
Culture tubes	BD Falcon	352059
LB Broth Miller	EMD	EM1.10285.0500
Nalgene Centrifuge Tubes	Thermo	3118-0050
Isoflurane	Butler Schein	NDC 11695-6776-1
Catheter needle 30G	Hamilton	91030
PE10 tubing	BD intramedic	427400
Von Frey Filaments	Stoelting	58025-31