Age-related changes in the innervation of the prostate gland

Abstract

The adult prostate gland grows and develops under hormonal control while its physiological functions are controlled by the autonomic nervous system. The prostate gland receives sympathetic input via the hypogastric nerve and parasympathetic input via the pelvic nerve. In addition, the hypogastric and pelvic nerves also provide sensory inputs to the gland. This review provides a summary of the innervation of the adult prostate gland and describes the changes which occur with age and disease. Growth and development of the prostate gland is age dependent as is the occurrence of both benign prostate disease and prostate cancer. In parallel, the activity and influence of both the sympathetic and parasympathetic nervous system changes with age. The influence of the sympathetic nervous system on benign prostatic hyperplasia is well documented and this review considers the possibility of a link between changes in autonomic innervation and prostate cancer progression.

The adult prostate gland grows and develops in an age-dependent manner. In aged males this development gives rise to abnormalities which are benign [benign prostatic hyperplasia (BPH)] and/or malignant (prostate cancer). Seemingly in parallel, autonomic nervous system activity changes in men with age, and this has been associated with diseases such as hypertension and BPH. This review describes the innervation of the prostate gland through the stages of adult life and explores the possibility that changes in autonomic nervous system activity may contribute to prostate cancer initiation and/or progression.

Fetal and Prepubescent Prostate Development

Morphogenesis of the human prostate gland occurs around the tenth week of gestation when circulating fetal androgen levels stimulate the differentiation of the endodermal urogenital sinus, causing the formation of solid epithelial outgrowths (prostatic buds).¹ The prostatic buds rapidly lengthen, arborize, cannulate and cytodifferentiate into basal and luminal epithelium.¹ The newly formed tubuloalveolar ducts grow and spread throughout the urogenital mesenchyme, which concurrently differentiates and matures into the smooth muscle-containing prostatic stroma. The growth and maturation of the tubuloalveolar ducts and stroma is dependant on androgens as well as the interaction between the urogenital mesenchyme and epithelial growths.¹ By the thirteenth week of gestation, there are approximately 70 primary ducts surrounding the developing urethra and by birth ductal branching is complete.¹ The pre-pubertal prostate is small, weighing approximately 2 g, and due to the low levels of testosterone, growth of the prostate during this period is limited.²

Prior to puberty, the prostate gland is quiescent and presumably not influenced by the autonomic nervous system. At the beginning of puberty, secretion of androgens from the testes cause the prostate to undergo a period of rapid development and growth ultimately reaching its full size and mature morphology by 18–20 y.²

The Young Adult Prostate Gland

The young adult prostate weighs approximately 20 g and is the largest of the male accessory reproductive organs. It is an alobular structure found posterior to the bladder that completely encapsulates the prostatic urethra and ejaculatory ducts.² The glandular elements of the prostate are made up of branching tubuloalveolar ducts with numerous secretory acini, surrounded by a thin fibromuscular stroma. The glandular elements or zones, which produce and drain prostatic secretions into the urethra, account for approximately 70% of the total prostate bulk with the fibromuscular stroma, comprising of connective tissue and smooth muscle, making up the remaining 30%.³

While testosterone is the primary circulating androgen produced by the testes, in peripheral tissues such as the prostate, testosterone is converted locally to dihydrotestosterone (DHT) by the action of the enzyme 5α-reductase.⁴ DHT is more potent than testosterone and has a higher affinity for the nuclear androgen receptor.⁵ Activation of the androgen receptor, via various mechanisms, results in cell proliferation and growth.¹ In addition to androgens, growth and proliferation of the prostatic stroma is mediated by estrogens, particularly estradiol acting at the ERα estrogen receptor.⁶ Estradiol is formed locally in the prostate from the conversion of testosterone by aromatase, which like 5α-reductase is localized primarily in the prostatic stroma.⁷ Furthermore, as with the development of the fetal prostate, reciprocal stroma-epithelial (mesenchyme-epithelial) interactions mediated by paracrine factors, in part under the influence of androgens and estrogens, play a vital role in the growth of the prostate.¹ Following the spike in androgen levels during puberty, circulating androgen levels stabilize around 20 y of age. Stabilization of androgen levels corresponds to a period of slow prostatic growth until approximately the age of 50.⁸

Innervation of the Adult Prostate Gland

Intact neuronal inputs and contractile mechanisms of prostatic smooth muscle are essential for the proper functioning of the prostate, as sympathetically mediated contractions of the prostatic smooth muscle expel prostatic fluid from the prostate into the ejaculate. The prostate is innervated by a rich supply of mixed autonomic postganglionic neurons that arise from the pelvic (inferior hypogastric) plexus, containing neuronal inputs from both sympathetic and parasympathetic neurons. The preganglionic sympathetic neurons arise from the lumbar spinal cord and descend to the pelvic plexus via the hypogastric nerve, whereas preganglionic parasympathetic neurons join the pelvic plexus from the pelvic nerve arising from the sacral spinal cord segment.^9,10

Consistent with the role of adrenergic nerves mediating contraction of the prostatic smooth muscle, the prostatic stroma is richly innervated with short noradrenergic nerves arising from the pelvic plexus that are absent from the prostatic glandular epithelium.¹¹ In the prostate, noradrenaline released from noradrenergic nerves activates G protein-coupled α₁-adrenoceptors, which results in smooth muscle contraction via an increase in intracellular calcium. In early in vitro contractile studies, prostatic smooth muscle contraction could be elicited by exogenously applied α-adrenoceptor agonists and such effects could be inhibited or blocked by non-specific α-adrenoceptor antagonists such as phentolamine (inhibits α-adrenoceptors but exhibits no selectivity toward α₁- or α₂-adrenoceptors)^12,13 or non-specific α₁-adrenoceptor antagonists such as prazosin (inhibits α₁-adrenoceptors but exhibits no selectivity toward α_1A-, α_1B or α_1D-adrenoceptor subtypes).¹⁴ As delineated by molecular and cloning studies, the α₁-adrenoceptor family consists of three subtypes: the α_1A-adrenoceptor, the α_1B-adrenoceptor and the α_1D-adrenoceptor.¹⁵ Experiments investigating mRNA expression and α₁-adrenoceptor density have indicated that the α_1A-adrenoceptor is the dominant subtype expressed in the prostate of various species, including humans, and that the α_1A-adrenoceptor is primarily localized to the prostatic stroma.^16,17 Pharmacological characterization studies have identified that a functional phenotype of the α_1A-adrenoceptor subtype, the α_1L-adrenoceptor, mediates the adrenergic contractile response in the human,^18,19 canine,²⁰ rabbit,²¹ guinea pig,²² rat²³ and mouse²⁴ prostates, with the α_1B-adrenoceptor and α_1D-adrenoceptor subtypes having little or no involvement in smooth muscle contraction.^19,25,26 Importantly, α_1L-adrenoceptors also mediate the contractile response to endogenously released noradrenaline in electrical field stimulation experiments.^19,27

In prostatic smooth muscle, stimulation of the G_q/11 protein-coupled α_1A-adrenoceptors (α_1L-adrenoceptors) results in the prototypical activation of phospholipase C and the subsequent production of the second messengers inositol 1,4,5-triphosphate and diacylglyercol. These in turn mediate smooth muscle contraction via activation of Ca²⁺-dependent and Rho kinase-dependent Ca²⁺-sensitization signaling pathways.²⁸ However, recent studies have shown that activation of c-jun N-terminal kinase²⁹ as well as phosphorylation of caldesmon³⁰ are involved in smooth muscle contraction of the prostate following α₁-adrenoceptor stimulation, indicating that further intracellular pathways are involved in α₁-adrenoceptor mediated contraction of the prostate.

In addition to α_1A-adrenoceptors, which directly mediate prostatic smooth muscle contraction, both α₂-adrenoceptors and β-adrenoceptors are also found in the prostate. The human prostate contains a population of α₂-adrenoceptors with a density comparable to³¹ or lesser than that of α₁-adrenoceptors^25,32 and are localized primarily in the glandular epithelium and vascular tissue with sparse stromal distribution.^32,33 Functionally, α₂-adrenoceptors reduce nerve-mediated contractions of the prostate via pre-junctional inhibition of neuronal noradrenaline release^19,31 and appear to be without a direct post-junctional role in contraction of the prostate.^25,34 Similarly, β-adrenoceptors have been found in the human, pig and rat prostates^35-37 and have been shown to inhibit α₁-adrenoceptor-mediated contractile responses in various experimental species as well as in the human prostate.^36,38-41 β-adrenoceptors inhibit α₁-adrenoceptor-mediated contraction via post-junctional activation of adenylate cyclase and accumulation of cyclic adenosine monophosphate (cAMP) resulting in relaxation of prostatic smooth muscle.³⁹ This response is most likely mediated by β₂-adrenoceptors; however β₁-adrenoceptors and β₃-adrenoceptors have also been implicated.^37,38,40 Recently, further interplay between α₁-adrenoceptors and β-adrenoceptors in the prostate has been proposed, as activation of α₁-adrenoceptors results in the phosphorylation of β-adrenoceptors, possibly via mechanisms involving G protein-coupled receptor kinases. Such an effect may enhance the α₁-adrenoceptor-mediated contractile response.⁴²

In addition to mediating contraction, adrenergic innervation plays a role in the growth of the prostate. In rodents, sympathectomy of the hypogastric nerve causes a reduction in prostatic weight in rats,⁴³ whereas chronic administration of α₁-adrenoceptor agonists induces proliferation and hyperplasia.^44,45 Multiple α₁-adrenoceptor antagonists have been shown to suppress stromal and epithelial cell growth in cell culture;⁴⁶ however this does not appear to be the result of specific blockade of the receptor and does not appear to translate to a clinical reduction in prostate volume.⁴⁷ In the rat, prostatic hyperplasia induced by phenylephrine activation of α₁-adrenoceptors has been strongly linked to activation of inflammatory pathways and the transforming growth factor β (TGFβ) signaling cascade.⁴⁵ Furthermore, human studies have shown that α₁-adrenoceptors in the prostate couple to non-contractile intracellular protein kinases involved in growth, proliferation and apoptosis,⁴⁸ indicating that activation of α₁-adrenoceptors in the human prostate couple to multiple pathways, some of which may be involved in proliferation and growth of the prostate.

Cholinergic innervation is found in both the stromal and glandular epithelial regions of the human,^11,49,50 guinea pig²⁷ and rat⁵¹ prostates. Responses elicited by cholinergic innervation in the prostate are mediated by G protein-coupled muscarinic receptors of which there are five subtypes. These are the M₁, M₃ and M₅ muscarinic receptors, which couple to G_q/11 proteins, and the M₂ and M₄ muscarinic receptors, which couple to G_i/o proteins.⁵² The endogenous agonist for all five subtypes is acetylcholine.⁵² In various species, muscarinic receptors are primarily localized in the glandular epithelium; however some muscarinic receptor expression is also found in the prostatic stroma suggesting a dual role in secretion and contraction. In agreement with the primary localization of muscarinic receptors on the glandular epithelium, stimulation of the cholinergic nerves or direct activation of muscarinic receptors results in the production of prostatic secretions.^53,54

Despite muscarinic receptors being located primarily in the glandular epithelium, they play a direct role in post-junctional contraction in the prostate, as in vitro contraction of isolated prostates elicited by electrical field stimulation or exogenous agonists can be inhibited by muscarinic receptor antagonists in the human,^12,31,55 canine,^41,56 rabbit,^57,58 guinea pig,^38,58,59 rat^58,59 and mouse^60,61 prostates. However, the magnitude of cholinergic contractions in the prostate is less than that observed for α₁-adrenoceptor stimulation.⁶² Additionally, muscarinic receptors regulate nerve mediated contractions by pre-junctional inhibition³¹ or facilitation⁶³ of neurotransmitter release. The muscarinic receptor subtype responsible for contraction differs between species. M₂ muscarinic receptors mediate contraction in the canine prostate⁵⁶ and are also the predominant subtype in human stromal cells where they inhibit the accumulation of cAMP.⁶⁴ In the guinea pig prostate, M₁ muscarinic receptors facilitate nerve mediated contraction,⁶³ whereas contraction of the rat and mouse prostates is mediated by M₃ muscarinic receptors,^61,65 which are localized on the prostatic smooth muscle.⁵¹

In the prostate, activation of muscarinic receptors would likely result in the activation of the prototypical signaling pathways which are phospholipase C for M₁, M₃ and M₅ and adenylate cyclase for M₂ and M₄ muscarinic receptors, respectively. Only a few studies have investigated the signaling pathways for the muscarinic receptor subtypes, which mediate prostatic contraction. Studies in cancer cell lines indicate that activation of the muscarinic receptors present in prostate smooth muscle can result in cAMP accumulation or phosphatidylinositol turnover.⁶⁴ However, recently, Rho kinase-dependent Ca²⁺ sensitization signaling pathways have been implicated in the muscarinic receptor-mediated contraction of the rat prostate.⁶⁶

As is the case in most physiological systems, the sympathetic and parasympathetic nervous systems appear to oppose each other in the form of a physiological balancing mechanism. This appears to be the case also in the control of prostatic growth as parasympathectomy in rats leads to increased prostate weight.⁴³

In addition to adrenergic and cholinergic innervation numerous other non-adrenergic, non-cholinergic neurotransmitters that can elicit or modulate contraction are found in the prostate. While these mechanisms have been shown to play a role in prostate contraction, their physiological role is uncertain and in general their contribution to physiological contraction is much less than that of noradrenaline acting at α_1A-adrenoceptors.

Adenosine 5′-triphosphate (ATP) is a known sympathetic co-transmitter with noradrenaline in rat and guinea pig prostates and mediates contraction via post-junctional P2X1 purinoceptors,^27,38,67,68 localized on the prostatic smooth muscle.^67-69 However, this effect seems to be species-dependent as ATP does not contribute to nerve mediated contractile responses in the mouse prostate^60,70 despite immunolocalization of P2X1-purinoceptors in the prostatic smooth muscle.⁷⁰ Following neuronal release, ectonucleotidases hydrolyse ATP to adenosine, which in turn can activate adenosine receptors. In the mouse prostate, pre-junctional A_2A adenosine receptors contribute to nerve mediated contraction by facilitating noradrenaline release⁷¹ whereas in the rat prostate, pre-junctional A₁ adenosine receptors inhibit excitatory neurotransmitter release.⁷² Furthermore, A₁ and A_2A adenosine receptors have a post-junctional role in contraction as the α₁-adrenoceptor-mediated response in cultured human prostatic stromal cells is enhanced or inhibited by A₁ or A_2A adenosine receptors via the inhibition and stimulation of adenylate cyclase, respectively.⁷³

Neuropeptide Y is co-localized in all adrenergic nerves in pelvic ganglia and is also found co-localized in some cholinergic nerves.¹⁰ Dense neuropeptide Y innervation is found throughout the prostatic stroma of humans,⁵⁰ guinea pigs, rats⁷⁴ and mice.^70,75 However, there are conflicting reports concerning the expression of neuropeptide Y receptors in the prostates of humans⁷⁶ and rats.⁷⁷ Curiously, despite dense innervation to the stroma and roles in modulating neurotransmitter release in other genitourinary tissues, most studies have found that neuropeptide Y does not appear to regulate or mediate contraction of the prostate^63,76,78 and may therefore instead be involved in prostate growth.⁷⁹ However, one study did find that in the human prostate, exogenous application of high concentrations of neuropeptide Y inhibits nerve-mediated contractions and relaxes noradrenaline pre-contracted prostate preparations.⁸⁰

In various species the prostate receives a rich supply of vasoactive intestinal polypeptide (VIP) containing nerves, which are co-localized predominantly in cholinergic nerves innervating the prostatic glandular epithelium.^10,11,75 Receptors for VIP have been found in the prostate;⁸¹ however due to their glandular distribution, VIP does not appear to be involved in contraction and instead may have roles in secretion or prostatic growth.⁸²

In contrast to neuropeptide Y and VIP, which have little if any role in the contraction of the prostate, other peptides are known to regulate or elicit prostatic contraction. The sensory neuropeptide calcitonin gene-related peptide (CGRP) has been found in nerves located within the stroma and/or glandular epithelium of numerous species^70,80,83-85 as well as neuroendocrine cells in the human prostate.⁸⁶ Functionally, exogenous CGRP has been shown to relax phenylephrine mediated contractions⁷⁸ or inhibit nerve mediated contractions⁸⁷ in the rat prostate. Furthermore, tachykinins also modulate prostate contraction. Neurokinin A and substance P have been found distributed sparsely in nerve fibers supplying the prostatic smooth muscle of human,⁵⁰ sheep,⁸⁸ canine,⁸⁵ guinea pig, rat^89,90 and mouse prostates.⁷⁰ Exogenous application of tachykinin agonists induces prostate contraction via neurokinin NK2 receptors in the human prostate,⁹¹ whereas neurokinin NK1 and NK2 receptors mediate contraction to exogenous agonists in cultured canine prostatic stromal cells.⁸⁵ Conversly, no direct contractile effects have been observed in the rat⁷⁸ or pig⁵⁸ prostates in response to substance P. In the guinea pig prostate, however, substance P and neurokinin A potentiate contractions elictited by nerve stimulation.⁹⁰

While not mediating contraction, nitrergic neuronal mechanisms regulate the tone of the prostatic smooth muscle by mediating relaxation. The prostate receives a dense innervation of nitric oxide synthase containing nerves, which are localized throughout the prostatic stroma and glandular epithelium.⁹² In addition, nitric oxide synthase is commonly co-localized in both noradrenergic and cholinergic nerves.^80,84 Electrical field stimulation relaxes in vitro prostate preparations that have been pre-contracted with noradrenaline in the human, canine,⁹³ rabbit^58,94 and guinea pig⁵⁸ tissue. This nerve-mediated relaxation can be blocked by inhibition of nitric oxide synthase or enhanced in the presence of nitric oxide donors.^58,93,94 The exogenous addition of nitric oxide donors also relaxes precontracted prostate tissues.^58,80,93,95 Furthermore, when the adrenergic and cholinergic components of nerve mediated contraction are blocked, electrical field stimulation relaxes prostatic smooth muscle, which can be inhibited or enhanced by nitric oxide synthase inhibitors or nitric oxide donors, respectively.⁹³ Phosphodiesterase (PDE) enzymes, which are involved in the hydrolysis of cyclic nucleotides produced by the action of nitric oxide, are also found in the human prostate.⁹⁵ Moreover, inhibitors of the PDE4 and PDE5 isoforms have been shown to relax prostatic tissues precontracted with noradrenaline or endothelin-1.⁹⁵

Age-Related Changes in Prostate Innervation

At approximately age 50, the steady growth of the prostate slowly accelerates.⁸ In parallel, a number of studies have shown an age-related decrease in the innervation to the prostate over this time.^50,96 At the same time, an increase in α_1A-adrenoceptor mRNA expression has been observed in aged human prostate.^97,98 This is in contrast to the aged rat prostate where α_1A-adrenoceptor mRNA expression is lower.⁹⁹ In the aged human prostate the contractile response to exogenously adminstered α₁-adrenoceptor agonists remains the same³⁶ or is increased.⁵⁵ However, in similar studies of the aged rat prostate, a decrease in α₁-adrenoceptor density⁹⁹ and distribution³³ was observed, which resulted in a reduced contractile response mediated by α₁-adrenoceptor agonists.⁹⁹ While the reason for the difference between species is unclear, it might be due to differences in age. Rats used in the previous studies were 18–22 mo of age, corresponding to an approximate human age of only 45–55 y old.¹⁰⁰

Research in our laboratory has also noted that prostates taken from 12-mo-old α_1A-adrenoceptor knockout mice are smaller than those taken from wild-type litter mate controls at the same age (Fig. 1). As previously mentioned, sympathetic innervation is known to play a role in the growth of the rat prostate, as surgical sympathectomy of the pelvic ganglia⁴³ reduces the size of the prostate, while sympathetic stimulation results in prostate growth.⁴⁴ Furthermore, in patients with spinal cord injury resulting in severe paralysis, smaller prostate size is observed.¹⁰¹ In rats, the in vivo administration of non-selective α₁-adrenoceptor antagonists results in a reduction in prostatic weight¹⁰² as well as cellular proliferation.¹⁰³ This effect appears to be dose dependent, as low doses of α₁-adrenoceptor antagonist have no effect on prostatic weight.¹⁰⁴ Our observations with knockout mice (Fig. 1) indicate that the α_1A-adrenoceptor subtype is not only responsible for the nerve mediated contractile response but in the mouse prostate is responsible for sympathetically mediated growth as well.

Figure 1. Aged α_1AKO mice. Comparison of mean mouse weight (A), isolated prostate weight (B) and mean contractile response to 80 mM KCl Krebs-Henseleit solution (C) from aged wild-type (WT, open columns) and α_1A-adrenoceptor knockout mice (α_1AKO, black columns). Columns represent mean weight/contractile force ± S.E.M., n = 10–14. p-values, **p < 0.01 vs. control, were calculated by an unpaired t-test and represent the probability of genotype affecting prostate weight in the aged mouse.

Cholinergic innervation of the human prostate was shown not to change with age.⁹⁶ Similarly, in the aged human prostate³¹ as well as the aged canine¹⁰⁵ and rabbit¹⁰⁶ prostates, no change in muscarinic receptor density was observed. Whereas, in the aged rat prostate, M_1–3 muscarinic receptor mRNA decreased¹⁰⁷ as did M₃ muscarinic receptor density.^65,107 However, studies using antibodies in the rat prostate, showed an increase in M₂ muscarinic receptors in the rat prostate with age.³³ Therefore the effect of age on the cholinergic innervation in the aged prostate appears to be highly subtype and species dependent. This is of note as a possible synergistic adrenergic-cholinergic action has previously been observed in the human prostate.³¹

Many inhibitory mechanisms of prostate contractility have also been studied with regard to aging. An increase in α₂-adrenoceptor density has been observed with age in the human,²⁵ rat³³ and rabbit¹⁰⁶ prostates. In the prostate, α₂-adrenoceptors are primarily responsible for inhibition of noradrenaline release and do not play a direct contractile role,¹⁷ therefore, the implications of this phenomenon are unknown and could be complex. Expression of β-adrenoceptors, which are capable of relaxing electrical field stimulation and agonist mediated contraction,¹⁷ have also been shown to be reduced in the aged rat prostate.³³ Furthermore, a decrease in the activity of adenylate cyclase activated by the β-adrenoceptor agonist isoproterenol occurs in the aged rat prostate.¹⁰⁸ In contrast, β-adrenoceptor expression does not change in the aged rabbit prostate.¹⁰⁶ Additionally, nitrergic innervation and nitric oxide mediated relaxation is decreased in the aged rabbit⁹⁴ and guinea pig¹⁰⁹ prostates. Therefore, a reduction in the inhibitory mechanisms of contraction in the prostate with age may play a role in the development of lower urinary tract symptoms (LUTS) associated with BPH.

BPH

BPH is a consequence of age- and androgen-dependant growth of the human prostate and affects approximately 50% of men by the age of 60 and 90% by 90 y of age.² BPH is a purely histological description of the prostate and should not be confused with LUTS associated with or secondary to BPH.¹¹⁰ While rarely life threatening, LUTS affect approximately 50% of men aged 50 to 80 y¹¹¹ and refers to a number of symptoms that can be found in either men or women that can severely affect their quality of life.¹¹² LUTS may be caused by BPH, however other factors such as detrusor over- or under-activity may also result in such symptoms.¹¹³ The LUTS can be categorized as related to urine storage, urine voiding and post-micturition. Urine storage symptoms relate to urgency and frequency of urination, nocturia and incontinence. Urine voiding-related symptoms include hesitancy, poor flow, intermittency and straining, while post-micturition symptoms include post-void dribble and a sense of incomplete emptying.¹¹⁰ Due to the high prevalence of LUTS accompanying BPH, there are significant costs associated with treatment that may be expected to rise with the aging population.¹¹⁴

BPH is characterized by a progressive nodular increase in the number of epithelial and stromal cells, which occurs initially in the transition and periurethral zones of the human prostate.¹¹⁵ Hyperplasia of the stoma is predominant and results in a greater proportion of smooth muscle relative to the glandular epithelium and an increase in the muscular tone of the hyperplasic prostate.¹¹⁶ BPH can lead to benign prostatic enlargement and in turn benign prostatic obstruction of the urethra. Benign prostatic obstruction is one cause of bladder outlet obstruction; others include bladder neck obstruction and urethral stricture,¹¹⁷ which increases bladder pressure and increases urethral resistance impairing the flow of urine from the bladder, leading to the voiding symptoms associated with LUTS.¹¹⁸ Bladder outlet obstruction due to BPH may also result in storage symptoms by inducing overactivity of the bladder detrusor.¹¹⁹

Effects of BPH on Prostate Innervation

In parallel with BPH development, α₁-adrenoceptor density in the human hyperplasic prostate is increased¹²⁰ or remains the same as the normal prostate.^25,36 Consequently there is an increase in the tone of the prostatic smooth muscle, which constricts the urethra, contributing to the LUTS associated with BPH. In the hyperplasic prostate the increase in tone of the prostate is mediated, in part, by neuronal noradrenaline acting at α_1L-adrenoceptors to cause smooth muscle contraction and this mechanism forms the basis for the treatment of BPH with α₁-adrenoceptor antagonists. However, prostatic smooth muscle contraction is also mediated by numerous other receptor systems,^74,121 such as acetylcholine acting at muscarinic receptors or ATP acting at purinoceptors. Therefore, blockade of these receptors, particularly muscarinic receptors, has been hypothesized as a suitable additional target for a better pharmacological treatment for BPH.¹²² Changes with age in these, or other mediators of prostatic smooth muscle contraction, may play a role in the development of LUTS associated with BPH by increasing prostatic tone.

Given the important role of α₁-adrenoceptors in the treatment of BPH, many studies have previously investigated the effect of age or disease on adrenergic contractile mechanisms, which demonstrated significant variation between species and experimental conditions. In BPH as with age, total innervation as well as adrenergic innervation of the human prostate gland decreases.^50,96 Adrenoceptor studies of the human prostate show contrasting effects. α_1A-adrenoceptor mRNA was increased in the diseased human prostate,⁹⁷ as was observed with age in normal human prostate.⁹⁸ At the receptor level, α₁-adrenoceptor density in the hyperplasic human prostate is increased¹²⁰ or remains the same as in normal prostate.^25,36 Studies of the contractile response in human prostate mirror those investigating α₁-adrenoceptor density. Whereby, the contractile response of human hyperplasic prostate to exogenously applied α₁-adrenoceptor agonists were shown to remain the same^31,36,55 or were increased³¹ compared with normal prostate.

In contrast to the adrenergic component of contraction, only a few studies have investigated the effect of age or disease on the cholinergic component of contraction in the prostate. As observed with the adrenergic component, previous studies show differences between species and experiments. Unlike in normal aged prostates, in hyperplasic prostates, acetylcholinesterase staining decreased.^49,50 However, as seen in the aged prostate, hyperplasic human prostate³¹ shows no change in muscarinic receptor density. Conversely, an upregulation of the M₃ muscarinic receptor, but not the M₁ or M₂ muscarinic receptor subtypes, was observed in the human hyperplasic prostate.¹²³ Finally, acetylcholine was shown to potentate the noradrenergically mediated contractile response in the hyperplasic human prostate,³¹ but not in un-diseased tissue.³¹ In general, the effect of BPH on the cholinergic contractile response in the prostate gland is still poorly understood.

Changes in inhibitory inputs of prostatic contraction have also been widely investigated with respect to BPH. Expression of β-adrenoceptors, which are capable of relaxing electrical field stimulation and agonist mediated contraction,¹⁷ were shown to be decreased in the hyperplasic human prostates.³⁶ Furthermore, a decrease in the activity of adenylate cyclase activated by the β-adrenoceptor agonist isoproterenol occurs in the hyperplasic human prostate.³⁶ These studies implicate a reduction in the β-adrenoceptor inhibitory mechanisms of contraction in the prostate as playing a role in the development of LUTS associated with BPH. In contrast, in the hyperplasic human prostate expression of α₂-adrenoceptors was shown to be increased.^25,31

Prostate Cancer

Approximately 20,000 men are diagnosed with localized (organ-confined) prostate cancer in Australia each year. In many patients the tumors are slow-growing and are not associated directly with mortality. Therefore these men can be successfully treated with conventional treatment regimes including surgical removal of the prostate (radical prostatectomy) or radiotherapy. If the patient relapses and the cancer returns, they are treated with androgen ablation/deprivation therapy, which can reduce the tumor and circulating prostate specific antigen (PSA) to undetectable levels. However, in most cases the cancer will eventually recur in an androgen independent/hormone refractory/castration resistant form that usually results in lethal bone metastasis.¹²⁴ Chemotherapeutic regimes for advanced prostate cancer are often still used but generally result in only a small increase in survival time.^125,126

The strongest risk factor for the development of prostate cancer in men is age. The chance of developing the disease rises rapidly after the age of 50 with 1 in 11 Australian men developing prostate cancer by the age of 70. The sympathetic nervous system is implicated in disorders of the aging male such as hypertension and BPH. In both of these age-related disorders, the sympathetic nervous system shows signs of overactivity and symptoms can be controlled by the use of therapeutic drugs which block the effects mediated by adrenoceptors (e.g., α₁-adrenoceptor antagonists, β-blockers).

In humans, α₁-adrenoceptor antagonists used in the treatment of BPH have been shown to induce prostate apoptosis.^98,103,127 This apoptotis inducing effect of α₁-adrenoceptor antagonists has also been shown in prostate cancer cell lines in vitro^46,128,129 and in vivo in mice bearing a tumor following subcutaneous xenograft injection of PC3¹²⁸ or LNCaP¹²⁹ cells. The antitumorigenic effect of the α₁-adrenoceptor antagonists was originally postulated to be due to antiangiogenic effects on the prostate vasculature;¹³⁰ however, their apoptotic effects on prostate cancer cell lines in culture^46,128 suggest an alternate mechanism of action. Nevertheless, several reports indicate that only the quinazoline based α₁-adrenoceptor antagonists have anti-apoptotic efficacy against prostate cancer cells and that this is by an α₁-adrenoceptor-independent mechanism.¹³⁰ However, in vivo data generated from our laboratory using α_1A adrenoceptor knockout mice indicates that there is also a mechanism which involves the sympathetic nervous system and more specifically α₁-adrenoceptors (Fig. 1). In support of this, it has previously been shown that activating the α₁-adrenoceptor signaling pathway induces proliferation of human prostatic stromal cells.¹³¹

β-adrenoceptors have also been implicated in metastasis development in models of prostate and other cancers. In vitro studies have shown that active migration of tumor cells can be induced by noradrenaline using cell lines derived from colon, breast or prostate cancer cells.¹³² Furthermore, this tumor cell migration could be inhibited by the β-adrenoceptor antagonists propranolol or ICI 118,551.¹³² Noradrenaline induced development of metastasis following in vivo xenograft of PC3 cells in BALB/c nude mice can also be inhibited by the β-blocker propranolol.¹³³ Indeed it has recently been shown that inducing stress in an in vivo mouse xenograft model of prostate cancer can artificially activate the sympathetic nervous system to stimulate β-adrenoceptor induced metastatic effects.¹³⁴ This metastatic effect was mediated by increased levels of circulating adrenaline and could be inhibited by β-adrenoceptor antagonists. With regard to parasympathetic cholinergic mechanisms, stimulation of muscarinic receptors of the M₃ subtype have been shown to stimulate proliferation of LNCaP prostate cancer cells as well as benign and cancerous primary prostate cells.¹³⁵ Similarly, in studies of the cancerous rat prostate, an increase in M₃ muscarinic receptor density was observed.¹³⁶ This suggests that M₃ muscarinic receptors may play a significant role in prostate cancer tumor growth and possibly also androgen-independent tumor progression.

It has previously been hypothesized that there is an association between increased sympathetic activity and prostate cancer,¹³⁷ and the scientific literature abounds with circumstantial evidence for the involvement of the sympathetic nervous system in prostate cancer progression. For instance obesity, which is associated with high sympathetic tone,¹³⁸ is also associated with increased prostate cancer risk,¹³⁹ as is cardiovascular disease.^140,141 In clinical studies on schizophrenia patients, there is a significant association of reduced risk of prostate cancer among patients treated with neuroleptic medication,¹⁴² particularly those who had been treated with high dose phenothiazine neuroleptics such as chlorpromazine.¹⁴² Although these drugs primarily target dopamine receptors, high doses of this class of neuroleptics are known to antagonize α₁-adrenoceptors, so it is notable that this relationship was seen only in patients treated with a high cumulative dose. Diabetes, on the other hand, which is associated with sympathetic neuropathy, is associated with a lower risk for late stage prostate cancer but no association in early stages.¹⁴³ Similarly, lower body sympathetic dysfunction due to spinal cord injury is associated with a low incidence of prostate cancer diagnosis.¹⁴⁴ Finally, the most striking clinical evidence comes from a study on antihypertensives which indicated that β-blockers and the long-term use of α-adrenoceptor antagonists may prevent prostate cancer whereas other classes of antihypertensives such as calcium channel blockers or angiotensin-converting enzyme inhibitors do not influence prostate cancer risk.¹⁴⁵

In clinical trials, a study conducted on 4,070 men in the U.S. found that men exposed to α₁-adrenoceptor antagonists have a 1.5 times lower risk of developing prostate cancer than unexposed men.¹⁴⁶ This study only looked at men taking the quinazoline based antagonists doxazosin, prazosin and terazosin and therefore observed effects may be due to non-adrenoceptor mechanisms. Conversely, evidence from a larger observational cohort study of 23,320 Finnish men showed that overall prostate cancer risk, while not reduced among α-adrenoceptor antagonist users compared with non users, showed a decreased incidence of high grade tumors,¹⁴⁷ indicating a decrease in tumor aggression. This second study looked at the effects of the widely used sulphonamide based α₁-adrenoceptor antagonist tamsulosin as well as the quinazoline based antagonist alfuzosin. The use of the non-quinazoline tamsulosin in this study is likely to be the cause of this novel effect on tumor aggression, which is likely to be α₁-adrenoceptor mediated. The evidence found in this second epidemiological study is consistent with the sympathetic nervous system, which is often overactive in the aging male, contributing to the progression of prostate cancer. The former study, on the other hand, is consistent with the previously reported antiproliferative α₁-adrenoceptor independent effects of the quinazoline-based compounds.

Interpretation of antitumorigenic effects in prostate cancer cell lines is difficult since the expression of the different adrenoceptor and muscarinic receptor subtypes in the different prostate cancer cell lines is not well characterized. In addition, these commonly studied cell lines are likely to be phenotypically different under different conditions in differerent laboratories. For example, based on single-cell RNA sequencing data, the β₂-adrenoceptor is expressed at significant levels in the PC3 and LNCaP prostate cancer cell lines while the M₃ muscarinic receptor is expressed only in PC3 cells¹⁴⁸ while the α_1A-adrenoceptor is not expressed in either of these cell lines. However, there are inconsistencies between the sequencing data and functional studies on α_1A-adrenoceptors and muscarinic receptors. Two papers have suggested a role for the α_1A-adrenoceptor in proliferation of LNCaP cells¹⁴⁹ and chemoresistance of DU145 cells;¹⁵⁰ however receptor detection was based on α_1A-drenoceptors antibodies, which are notorious for lack of specificity.¹⁵¹ The muscarinic receptor agonist carbachol likewise stimulates proliferation of LNCaP cells despite an apparent lack of mRNA for any of the relevant acetylcholine receptor subtypes.¹³⁵

Conclusions

Prostate disease and autonomic nervous system activity appear to change with age in a parallel and similar manner. Further research into whether this association is coincidental or not may elucidate the physiological mechanisms involved in clinical observations such as those that show prostate cancer patients who have been exposed to α₁-adrenoceptor antagonists for the treatment of BPH have a lower incidence of high grade tumors.¹⁴⁷ Validation of such a physiological mechanism will indicate a novel molecular target for chemotherapy of metastatic androgen-independent prostate cancer. This will in turn drive the development of new treatment strategies for these forms of currently treatment resistant prostate cancers. In addition, α₁-adrenoceptor antagonists, β-blockers and muscarinic receptor antagonists are already currently available, reasonably well-tolerated and very effective in the treatment of LUTS and hypertension. Understanding these signaling pathways may provide evidence for the immediate use of such a treatment strategy by clinicians to produce a survival advantage for advanced prostate cancer patients.

Glossary

Abbreviations:

ATP

adenosine 5′-triphosphate

BPH

benign prostatic hyperplasia

cAMP

cyclic adenosine monophosphate

CGRP

calcitonin gene-related peptide

DHT

dihydrotestosterone

LUTS

lower urinary tract symptoms

PDE

phosphodiesterase

PSA

prostate specific antigen

TGFβ

transforming growth factor β

VIP

vasoactive intestinal polypeptide

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.