**1. Introduction**

Chronic pelvic pain (CPP) is chronic or persistent pain perceived in pelvic structures for more than six months with continuous or recurrent pelvic pain as well as with symptoms suggestive of the lower urinary tract, sexual, bowel, pelvic floor or gynecological dysfunction in men and women [1]. CPP may be divided into situations with well-defined classical pathology (such as infection or cancer) and those without confirmed etiology [1]. According to this classification, the former is considered to be "specific disease-associated pelvic pain," and the latter is described as "chronic pelvic pain syndrome" (CPPS) [1]. The incidence rate of CPP in women is around 6% to 27% worldwide [2]. CPP in women is reported to affect 18–50-year-old women, mostly [3].

CPPS is a subdivision of CPP and the pain syndrome may be focused within a single organ or more than one pelvic organ, including bladder, urethra, vagina, rectum, anus and whole pelvic musculatures [1]. When the pain is localized to a single organ, some specialists use an end organ term such as Bladder Pain Syndrome (BPS), and use "syndrome" to indicate pain localized in more than one organ site [1]. However, some specialists sub-divide pelvic pain through psychological and functional symptoms rather than anatomy [1].

CPPS of women can exhibit different symptoms, including dyspareunia, dysmenorrhea, dyschezia, and non-menstrual pelvic pain. It may also cause lower urinary tract symptoms, such as frequency, urgency, difficulty urinating, and pain with urination. The most important evaluations for diagnosis of CPPS are medical history and physical examinations. [4]. Vaginal examination is critical to diagnose pelvic floor spasm due to the easily palpable taut muscles. Electromyography, perineometry, vaginal manometry, and digital assessment of pelvic floor muscle (PFM) are also conducted for diagnosis [5]. There are limited laboratory testing and imaging for diagnosis, and it should be considered to be evaluated by laparoscopic or urologic evaluation according to the clinical findings [4].

There is no single definitive etiology or standard management for CPPS. It is supposed that CPPS results from a combination of risk factors, such as neurological, mechanical, and biochemical factors. More than one-half of CPP patients live with interstitial cystitis/painful bladder syndrome (IC/BPS), endometriosis, irritable bowel syndrome, or pelvic adhesions [6,7]. Spasticity of the PFMs, which leads to an increase in muscle tone, has been proposed to play one of the important pathogenic factors of CPPS [8]. PFMs spasm may result from a primary event on the pelvic floor musculatures or secondary to other diseases related to psychological or pathological disorders. It is another significant problem that may decrease quality of life and increase health care costs.

The European Association of Urology (EAU) proposed comprehensive guidelines about the diagnosis and management of CPPS [1]. Management for CPPS remains limited, and the treatment points are usually symptom relief [9]. Secondary disease processes associated with CPPS in women should receive targeted treatment first. The etiologies of CPPS involve multiple mechanisms, so treatments for CPPS need a holistic approach including behavioral, physical, psychological, and sexual components. Modalities of treatments for CPP involve behavioral interventions, physical therapy, medications, surgical interventions, and alternative therapies [4]. The EAU guidelines suggest simple analgesics, such as nonsteroidal anti-inflammatory drugs, to be the first-line therapy of general management. Opioids and neuropathic analgesics, such as tricyclic antidepressants, anticonvulsants, and gabapentin, should be used as further medications if simple analgesics fail. If medications fail to provide symptom relief, nonpharmacologic managements, such as nerve blocks, suprapubic transcutaneous electrical nerve stimulation, sacral neuromodulation, and injection with Botulinum toxin A (BoNT-A) should be considered to help treat CPPS.

Botulinum neurotoxin (BoNT) comprises seven different serotypes (A to G) and more than 40 subtypes [10]. These serotypes have a similar mechanism to inhibit the release of acetylcholine (ACh), but they have different potency of actions. Among different subtypes of BoNT-A, A1 is the only isoform for the therapeutic purpose because of the high potency and long duration of paralysis [11]. The only subtypes of BoNT for clinical purposes are A1 and B1 [12]. However, the paralytic effect of BoNT-B is not as efficient as the function of BoNT-A according to the results of Sloop et al. [13]. The most studied BoNT for clinical treatment is BoNT-A.

BoNT-A injection can reduce spasms and pressure in the PFMs of women with CPPS [14]. Many women with CPPS reported diminution of pelvic pain symptoms after BoNT-A injection. Reduction in pelvic pain can improve quality of life, social activity, working performance, sexual relationships, urinating pain, and mood situations [15]. BoNT-A injected into the pelvic floor musculature of women with CPPS has been reported to improve chronic pelvic pain symptoms and spasms of PFMs [14]. The organ specificity of pelvic pain syndrome in CPPS includes urology, gynecology, gastroenterology, neurology, sexology, and the pelvic floor [1]. This review will focus on the BoNT-A treatment in female CPPS, especially in PFMs pain, IC/BPS, and sexual pain syndrome.

#### **2. Mechanisms of BoNT-A for Treating CPPS**

Bacterium *Clostridium botulinum* was first isolated in 1895. BoNT-A was first purified from bacterium *Clostridium botulinum* in 1928 and its off-label use started in the 70s [16].

There are three major mechanisms of BoNT-A that function on the muscles, neural system, and inflammation to relieve pain symptoms [17]. BoNT-A plays an important function in the reduction of pain symptoms. It is believed that spasms and tenderness of the PFMs are highly associated with CPPS in women [18,19]. BoNT-A injection has been used to paralyze muscles, and its effect is localized, partial, and reversible. After injecting to the PFMs, BoNT-A can reduce the hypertonic pressure and improve pelvic muscle spasms.

BoNT-A is a selective neurotoxin that acts on neuromusculatures. After binding to terminal receptors on the motor neuron, it can inhibit the release of ACh to cause muscle paralysis. BoNT-A inhibits ACh vesicles releasing to the synaptic cleft by cleaving particular proteins, such as SNAP-25 or VAMP, which are essential for binding with ACh vesicles at the presynaptic membrane. Due to the effect of BoNT-A, there is no release of ACh in the synaptic cleft, and it can paralyze the innervated muscles subsequently [20]. This mechanism has been used to relieve the storage of lower urinary tract symptoms of IC/BPS such as frequency and urgency.

Animal studies reported that BoNT-A could inhibit the delivery of several neurotransmitters, such as calcitonin gene-related peptide (CGRP), glutamate, adenosine triphosphate (ATP), and substance P [21–25].

BoNT-A may block these neurotransmitters from releasing muscular nociceptors, and reduce the symptom of muscle pain in patients with CPPS [26]. BoNT-A could also inhibit the contraction of muscles via alpha and gamma motor neurons and block spasms of pelvic floor musculature, which results in relieving the pelvic pain caused by muscle spasms [26]. Current literature has shown that the use of BoNT-A can reduce the hypertonicity of PFMs to improve pain scores from CPPS patients.

In addition, BoNT-A has the analgesic effect of relieving pain symptoms. From the animal and human studies, increased expression of cell membranes receptors, such as the TRPV1 and P2 × 3, in the nociceptors may up-regulate the symptoms of neuralgia [27,28]. BoNT-A has been reported to reduce the expression of TRPV1 in rats with neuropathic pain [29].

After the injection of BoNT-A, paralysis of muscle occurs after 2–5 days [5]. The functional effects can typically last from three to six months [14]. The clinical efficacy of BoNT-A injection for CPPS in women was durable to 24 weeks [30]. This long-term but reversible effect has made BoNT-A an important therapy for a wide variety of neuromuscular diseases.

After formation of antibodies against BoNT-A, the duration of the BoNT-A effect and the therapeutic extent of the maximal treatment effect are usually reduced after a few BoNT-A applications (partial therapy failure) before complete therapy failure occurs [5].
